The  Collaborative Studies on the Genetics of Alcoholism  ( COGA ) is an eleven-center research project in the United States designed to understand the genetic basis of  alcoholism . [ 1 ]   Research is conducted at  University of Connecticut ,  Indiana University ,  University of Iowa ,  SUNY Downstate Medical Center ,  Washington University in St. Louis ,  University of California at San Diego ,  Rutgers University ,  University of Texas Health Science Center at San Antonio ,  Virginia Commonwealth University ,  Icahn School of Medicine at Mount Sinai , and  Howard University .


Henri Begleiter  and Theodore Reich were founding PI and Co-PI of COGA. Since 1991, COGA has interviewed more than 17,000 members of more than 2,200 families from around the United States, many of whom have been longitudinally assessed. Family members, including adults, children, and adolescents, have been carefully characterized across a variety of domains, including other  alcohol  and other substance-related phenotypes, co-occurring disorders (e.g., depression), electrophysiology, key precursor behavioral phenotypes (e.g., conduct disorder), and environmental risk factors (e.g., stress). This has provided us with a very rich phenotypic dataset to complement the large repository of cell lines and DNA for current and future studies. We have made this dataset widely available to advance the field: hundreds of researchers have worked with data generated as part of COGA through a variety of different mechanisms including data sharing through dbGaP and the Genetic Analysis Workshops, as COGA collaborators, through meta-analysis consortia including the Psychiatric Genomics Consortium, and as independent requestors for COGA samples and data. [ 2 ]


In studying alcoholism, COGA hopes to find better ways of treating alcoholism and improving the lives of the millions of people who suffer from alcoholism. The COGA project has achieved national and international acclaim for its accomplishments, and numerous articles about the study have been published in scientific journals. [ 3 ]  This project is funded by the federal government (National Institute on Alcohol Abuse and Alcoholism) and is one of the largest of its kind to be done in the United States.


COGA's aim is to identify the genes involved in  alcoholism .  There is a large body of twin-studies and adoption-studies that show that the risk for alcoholism has a genetic component.  COGA is trying to determine more specifically what genes are involved. There is no one gene that confers much risk for alcoholism, rather, thousands of variations in many genes contribute to the risk, as do many aspects of the environment. Examples of risk factors include one's level of response to alcohol, a person's neuroelectrochemistry, and other psychiatric disorders such as  Antisocial Personality Disorder  or  clinical depression .


COGA researchers will interview subjects using the Semi-Structured Assessment for the Genetics of Alcoholism (SSAGA), [ 4 ] [ 5 ]  specifically created for the COGA project.  Each subject is asked to participate in the SSAGA, with different versions for adolescents and for parents being interviewed about their children.  The SSAGA is a polydiagnostic psychiatric interview that will cover any drug or alcohol use as well as any emotional and/or medical problems the subject may have experienced.  The SSAGA is designed to use diagnostic criterion from the DSM III-R, DSM IV, and ICD-10. The SSAGA has been translated into nine languages and has been used in over 275 studies; researchers can use the SSAGA in their own projects. [ 6 ]  The wide adoption of the SSAGA ensures that data from COGA families are highly compatible with data from other studies and allows COGA to participate in numerous consortia that use meta-analytic methods to combine data and results.


COGA requests that their subjects provide a blood sample, which is then processed and examined at Rutgers University.  DNA is used to identify variations in genes that affect the risk for alcoholism and related disorders. [ 7 ]


The brain wave is called an  event-related potential  (ERP) and is similar to an EEG.  Brainwaves are monitored in a non-invasive procedure that involves placing a cap on the subject's head, similar to a bathing cap.  This cap will monitor the brain's natural electrical activity and will record the brain's response when the subject is presented with various stimuli.  Subjects respond to computer programs on a screen in front of them that flash various stimuli on the screen.


After a significant event, or an important stimuli to which the subject is instructed to respond, there is an increase in electrical activity.  COGA is investigating, for example, the P3 (or P300) peak of the ERP that occurs approximately 300 ms after the stimuli presentation.  Varying amplitudes of the wave have different behavioral implications.  The amplitude is fairly consistent throughout families and steady across time, suggesting an inherited component, as opposed to an environmental component.


The  Consortium for the Barcode of Life  ( CBOL ) was an international initiative dedicated to supporting the development of  DNA barcoding  as a global standard for species identification. [ 1 ]   CBOL's Secretariat Office is hosted by the National Museum of Natural History, Smithsonian Institution, in Washington, DC.  Barcoding was proposed in 2003 by Prof. Paul Hebert of the University of Guelph in Ontario as a way of distinguishing and identifying species with a short standardized gene sequence.  Hebert proposed the 658 bases of the Folmer region of the mitochondrial gene cytochrome-C oxidase-1 as the standard barcode region.  Hebert is the Director of the Biodiversity Institute of Ontario, the Canadian Centre for DNA Barcoding, and the International Barcode of Life Project (iBOL), all headquartered at the University of Guelph.  The  Barcode of Life Data Systems  (BOLD) is also located at the University of Guelph.


CBOL was created in May 2004 with support of the  Alfred P. Sloan Foundation , following two meetings in 2003, also funded by the Sloan Foundation, at the Banbury Center, Cold Spring Harbor Laboratory.  Since then, more than 200 organizations from more than 50 countries have joined CBOL and agreed to put their barcode data in a public database.  CBOL promotes DNA barcoding through workshops, working groups, international conferences, outreach meetings to developing countries, planning meetings for barcoding projects, and production of outreach material to raise awareness of barcoding.  CBOL's Database Working Group developed the data standard that  GenBank , the  European Bioinformatics Institute , and the  DNA Data Bank of Japan  have endorsed.  CBOL's Plant Working Group proposed matK and rbcL as the standard barcode regions for land plants; CBOL approved this proposal in late 2005.  The Fungal Working Group has identified ITS as the best barcode region for fungi, and CBOL's Protist Working Group is analyzing candidate regions for protistan groups. [ 2 ]  CBOL helped to plan and launch the global campaigns to barcode all species of fish and birds, and socioeconomically important groups like fruitflies.


One of CBOL's primary contributions to the success of barcoding was its outreach efforts to government agencies (agriculture, environment, conservation, and others) and international organizations ( CITES ,  Convention on Biological Diversity ,  Food and Agriculture Organization ) that could benefit from barcoding.


This  Smithsonian Institution  article is a  stub . You can help Wikipedia by  expanding it .


The  Council for Responsible Genetics  (CRG) was a  nonprofit   NGO  with a focus on  biotechnology .


The Council for Responsible Genetics was founded in 1983 in  Cambridge, Massachusetts .


An early voice concerned about the social and ethical implications of  modern genetic technologies, CRG organized a 1985 Congressional Briefing and a 1986 panel of the  American Association for the Advancement of Science , both focusing on the potential dangers of genetically engineered biological weapons. [ 1 ]   Francis Boyle  was asked to draft legislation setting limits on the use of genetic engineering, leading to the  Biological Weapons Anti-Terrorism Act of 1989 . [ 2 ]


CRG was the first organization to advance a comprehensive, scientifically based position against  human germline engineering . [ 3 ]  It was also the first to compile documented cases of  genetic discrimination , laying the intellectual groundwork for the  Genetic Information Nondiscrimination Act  of 2008 (GINA). [ 1 ]


The organization created both a Genetic Bill of Rights and a Citizen's Guide to Genetically Modified Food. Also notable are CRG's support for the "Safe Seeds Campaign" (for avoiding  gene flow  from  genetically engineered  to non-GE seed) and the organization of a US conference on Forensic DNA Databanks and Racial Disparities in the Criminal Justice System. [ 1 ]  In 2010 CRG led a successful campaign to roll back a controversial student  genetic testing  program at the  University of California, Berkeley . [ 4 ]   In 2011, CRG led a campaign to successfully enact [CalGINA] in California, which extended  genetic privacy  and nondiscrimination protections to life, disability and long term care insurance, mortgages, lending and other areas. [ citation needed ]


CRG issued five anthologies of commentaries:


CRG "fosters public debate about the social, ethical and environmental implications of genetic technologies." They list three central  principles : [ 1 ]


In 2007, CRG hosted a retreat to refresh the mission statement and determine goals for the future of the organization. The outcome was that CRG should:


The pioneering contributions of CRG to public interest initiatives concerned with appropriate use of biotechnologies are recounted in the book  Biotech Juggernaut: Hope, Hype, and Hidden Agendas of Entrepreneurial Bioscience  (Routledge, 2019).


The CRG publishes  Genewatch , [ 5 ]  America's first and (according to CRG in 2009) only magazine dedicated to monitoring biotechnology's social, ethical and environmental consequences. The publication covers a broad spectrum of issues, from  genetically modified food  to  biological weapons , genetic  privacy  and discrimination,  reproductive technology , and  human cloning . [ 1 ]  Established in 1983, [ 6 ]  the publication won the  Utne  Independent Press Award for General Excellence in the category of newsletters in 2006. [ 7 ]


A major source of CRG's funding is the  Ford Foundation , which provided $420,000 in grants during 2005-2007. [ 8 ]


The  European Genetics Foundation (EGF)  is a non-profit organization, dedicated to the training of young  geneticists  active in medicine, to continuing education in genetics/genomics and to the promotion of public understanding of genetics. Its main office is located in  Bologna, Italy .


In 1988 Prof. Giovanni Romeo, President of the European Genetics Foundation (EGF) and professor of Medical Genetics at the  University of Bologna  and Prof.  Victor A. McKusick  founded together the European School of Genetic Medicine (ESGM).


Since that time ESGM has taught genetics to postgraduate students (young  M.D.  and  PhD ) from some 70 countries.  Most of the courses are presented at ESGM's Main Training Center (MTC) in Bertinoro di Romagna (Italy), and are also available via  webcast  at authorized Remote Training Centers (RTC) in various countries in Europe and the Mediterranean area (Hybrid Courses). In the  Netherlands  and  Switzerland , medical geneticists must attend at least one ESGM course before admission to their Board examinations.


For these reasons, the School has been able to expand and to obtain funding from the  European Commission  and from other international organizations.


The European School of Genetic Medicine was founded in 1988 and saw rapid success, which necessitated that an administrative body be formed.  To this end the  European Genetics Foundation  was born in  Genoa  on 20 November 1995, with the following aims:


The ESGM began receiving funding from the European Union and from other international organizations including the  European Society of Human Genetics , the  Federation of European Biochemical Societies , the  March of Dimes  and  UNESCO .  Since its founding, the ESGM has become a model for other European institutions responsible for advanced training in genetics.


In order to fully realize all of the European Genetics Foundation's aims, the Foundation has identified a permanent site for the ESGM at the  Hermitage  of  Ronzano , a monastery situated in the hills of Bologna, at 3.5 km from the city center and 7 km from the airport.  The monastic order of the Servants of Maria, founded in Florence in 1233 and owner of the Hermitage, donated 16,000 m 2  for the purpose of constructing this center, which will be named the  Giuseppe Levi and Victor A. Mckusick Euro-Mediterranean Center for Genetics and Medicine .   Giuseppe Levi , a scientist, trained three Nobel Laureates ( Salvador Luria ,  Renato Dulbecco  and  Rita Levi-Montalcini ), and Victor A. McKusick is considered the father of  medical genetics  throughout the world.


The European Genetics Foundation envisions the Ronzano Center as a venue for advanced training, and as a think tank where experts in genetics and related disciplines from all over the world can discuss issues of scientific, social, and political relevance, and to develop problem-solving strategies to address these issues.


The new center will host:


Genetic Alliance  is a nonprofit organization, founded in 1986 by Joan O. Weiss, working with  Victor A. McKusick , to advocate for health benefits in the accelerating field of  genomic  research. This organization is a network of over 1,000 disease advocacy organizations, universities, government organizations, private companies, and public policy organizations. They aim to advance  genetic  research agendas toward health benefit by engaging a broad range of stakeholders, including healthcare providers, researchers, industry professionals, public policy leaders, as well as individuals, families and communities.  They create programs using a collaborative approach, and aim to increase efficiency and reduce obstacles in genetic research, while ensuring that voices from the involved disease communities are heard.  They also promote public policies to advance healthcare. Genetic Alliance provides technical support and informational resources to guide disease-specific  advocacy organizations  in being their own research advocates. They also maintain a biobank as a central storage facility for several organizations who otherwise would not have the infrastructure to maintain their own repository.


The organization was founded in 1986 by Joan O. Weiss, working with  Victor A. McKusick , to advocate for health benefits in the accelerating field of  genomic  research. [ citation needed ]  The organisation is an umbrella organisation for a number of charities  [ who? ]  dealing with genetic diseases. The founding chair of the Genetic Alliance in 1989 was  Ann Mercy Hunt  who had founded the  Tuberous Sclerosis Association . [ 1 ]


Genetic Alliance manages a  biobank , the Genetic Alliance Registry and Biobank (GARB), which is a cooperatively-managed clinical data and tissue sample repository.  GARB was established in 2003, [ 2 ]  and combines its standardized infrastructure with the interests and motivations of disease advocacy groups to develop effective diagnostic tools and treatments, to “accelerate the research enterprise beyond its usual pace and involving new players, the advocates.” [ 3 ]   Individual disease advocacy organizations maintain control over their data and samples throughout the research process, while enabling collaboration among scientists by leveraging their resources: well-characterized, robust data sets including pooled samples from people with the disease. [ citation needed ]


Participating organizations store their data and samples in GARB for on-going use in research, with the goal of developing new diagnostic tools and treatments for diseases. [ 3 ]  Genetic Alliance provides the facility, collection, processing and archiving of samples stored in the biobank, as well as training and tools for organizations to effectively participate in this resource. GARB is governed collaboratively by an advisory board, with representatives from the seven currently participating organizations, which determines policies regarding stewardship, protection and usage of the data. [ 2 ]  GARB has an  institutional review board  (IRB), which approves data collection protocols, material transfer agreements, and documents used in informed consent and minor assents. [ citation needed ]


Organizations participating in GARB pay an annual fee to have access to GARB's IRB-approved protocols, training and technical assistance, and infrastructure for tissue banking. Stored data include “DNA, RNA, cell lines, tissue, organs, self-reported data, medical images, medical records,” [ 2 ]  amounting to over 30,000 samples and clinical records. Researchers can apply for access to samples and data in GARB via the advocacy organization responsible for them. [ citation needed ]


GARB was based on a  biobank  established by PXE International in 1995.  PXE International is a disease  advocacy organization , founded by Sharon Terry and Patrick Terry. In managing their biobank, they developed a series of protocols and systems for data collection, storage and use. Their efforts contributed to significant scientific advances, including the discovery of the  gene  responsible for  Pseudoxanthoma elasticum  (PXE). Drawing from their experiences, they began mentoring other disease advocacy organizations in developing new biobanks. [ 3 ]  The founders of the PXE International biobank established GARB to provide a new model for facilitating research, “on the principal that a shared infrastructure would facilitate easy flow of resources and accelerate disease-specific research.” [ 4 ]   A central value upon which GARB was established is a respect for specimen donors as participants in a dynamic community-engaged process.  Samples and data may be reused in many research studies, and re-contacting participants is possible to ensure on-going interaction and communication with those who donate to GARB. [ 2 ]


The  Genetic Discrimination Observatory  ( GDO ) is a  Montreal -based international network of researchers and other stakeholders who support the research and prevention of  genetic discrimination  (GD)—discrimination based on genetics or other predictive health information. [ 1 ] [ 2 ] [ 3 ] [ 4 ]  Their headquarters are currently located at the Centre of Genomics and Policy at  McGill University  in Montreal.


The GDO staff comes from various fields such as  genetics ,  ethics ,  law ,  sociology , and  public policy . [ 5 ] [ 6 ] [ 7 ]  The GDO received initial funding from  Genome Canada  ( Génome Québec  in French), the  Fonds de recherche du Québec – Santé , and the Network of Applied Medical Genetics (RMGA). [ 8 ]


In 2018, the GDO initiated its first project, the " Forum québécois sur la discrimination génétique " (Quebec forum on genetic discrimination) in  Quebec . [ 9 ] [ 10 ] [ 11 ]


The GDO provides information about different countries and areas using interactive world views that show studies related to genetic discrimination and other categories in a live map format on the GDO's website. [ 12 ] [ 13 ] [ 14 ]  It is planned to cover nineteen different jurisdictions. [ 2 ]


The GDO provides individuals from Canada, Mexico, the United Kingdom and the  United States  the opportunity to report specific cases of genetic discrimination or health-based discrimination confidentially. [ 15 ] [ 2 ]


Germplasm Resources Information Network  or  GRIN  is an online  USDA   National Genetic Resources Program  software project to comprehensively manage the computer database for the holdings of all plant  germplasm  collected by the  National Plant Germplasm System . [ 1 ]


GRIN has extended its role to manage information on the germplasm reposits of insect ( invertebrate ),  microbial , and animal species (see  sub-projects ). [ 2 ]


The site is a resource for identifying  taxonomic  information (scientific names) as well as common names [ 3 ]  on more than 500,000 accessions (distinct  varieties ,  cultivars  etc.) of plants covering 10,000 species; [ 4 ] [ 5 ]  both economically important ones [ 3 ]  and wild species. It profiles plants that are invasive or noxious weeds, [ 3 ]  threatened or endangered, [ 3 ]  giving out data on worldwide distribution [ 3 ]  of its habitat; as well as passport information. [ 6 ]  GRIN also incorporates an Economic Plants Database. [ 3 ]


The network is maintained by GRIN's Database Management Unit (GRIN/DBMU). [ 2 ]  GRIN is under the oversight of  National Germplasm Resources Laboratory (NGRL) in  Beltsville, Maryland , [ 6 ]  which in 1990 replaced its forerunner, the Germplasm Services Laboratory (GSL), [ 2 ]  that had formerly run GRIN. [ 7 ]  Since November, 2015 GRIN has been running on GRIN-Global software produced by a collaborative project between the USDA and the  Global Crop Diversity Trust . [ 8 ]


A stated mission of GRIN is to support the following projects: [ 9 ]


Human Genetics Alert  (abbreviated  HGA ) is a secular, independent  watchdog group  based in  London ,  England . It advocates against uses of  reproductive technology  and  human genetics  research that it considers harmful. [ 1 ]


Human Genetics Alert was founded in 1999 as the  Campaign Against Human Genetic Engineering . It was reborn as Human Genetics Alert (HGA) in December 2000, at which point it began to receive funding from the  Joseph Rowntree Charitable Trust . [ 1 ]  HGA stopped receiving funding from the Joseph Rowntree Charitable Trust in 2005. [ 2 ]  As of December 2018, its director was David King. [ 3 ]


Human Genetics Alert states that it is not opposed to all genetics research, but that it is opposed to certain proposed uses of it, including  human genetic engineering ,  human cloning , and  genetic discrimination . [ 4 ]  The organization has also opposed prenatal  sex selection [ 5 ]  and the proposal that led to the establishment of the  UK Biobank . [ 6 ]


The  Human Genetics Commission  (HGC) was an advisory  non-departmental public body  that advised the  UK government  on the ethical and social aspects of  genetics . This included  genetic testing , cloning and other aspects of molecular medicine. The Commission was created after a review of the UK government biotechnology advisory framework in 1999.  It was chaired initially by the lawyer,  Baroness Helena Kennedy QC  and, from 2007 to 2009, the acting chair was  Sir John Sulston . From 2009, the Commission was chaired by Professor Jonathan Montgomery and comprised 21 members whose backgrounds include the  law ,  medicine , consumer affairs, philosophy and ethics, scientific research, and clinical practice. Representatives of the Chief Medical Officers of  England ,  Scotland ,  Wales , and  Northern Ireland  also sat on the Commission. [ 1 ] [ 2 ]


The Commission was abolished when  quangos  were reviewed by the newly elected government in October 2010. The Commission published its final paper in May 2012. [ 1 ] [ 3 ] [ 4 ]


Human Longevity, Inc.  is a  San Diego –based venture launched by  Craig Venter ,  Robert Hariri  and  Peter Diamandis  in 2013. Its goal is to build the world's most comprehensive database on human genotypes and phenotypes, and then subject it to machine learning so that it can help develop new ways to fight  diseases associated with aging . [ 1 ]  The company received  US$80 million  in investments in its Series A offering in summer 2014 and announced a further $220 million  Series B  investment offering in April 2016. [ 2 ]  It has made deals with drug companies  Celgene  and  AstraZeneca  to collaborate in its research. [ citation needed ]


While it is conducting research, the company is offering a wellness service known as "Health Nucleus," which offers customers a range of medical tests such as a full genome sequencing and tests for early indications of  cancers ,  Alzheimer's  and  heart disease . [ 3 ]  This testing is meant to help people catch diseases earlier than otherwise possible and to identify risk factors for diseases later in life. [ 4 ]


At the start of 2017, the company hired Cynthia Collins from GE Healthcare, and Venter became Executive Chair. The company's chief operating officer, Mark Winham, left the company in mid-2017, and Collins and the company's chief medical officer, Brad Perkins, left in December. Venter stepped back into the  CEO  role, but announced in May 2018 that he was leaving the company to return to the J. Craig Venter Institute. [ 5 ]  Venter was sued for allegedly 'stealing trade secrets' at Human Longevity. [ 6 ]  The case has been dismissed. [ 7 ]


In November 2019, Human Longevity closed $30 million financing from Emerging Technology Partners (ETP) and other leading healthcare investors to renew its commitment to longevity and precision health. [ 8 ]


In November 2019,  C. Thomas Caskey , professor of molecular & human genetics at the  Baylor College of Medicine , joined the company as Chief Medical Officer. [ 9 ]


In January 2020, Human Longevity published a study in the journal  Proceedings of the National Academy of Sciences (PNAS)  that showed that by integrating whole-genome sequencing with advanced imaging and blood metabolites, clinicians identified adults at risk for key health conditions. [ 10 ]


In June 2020, Human Longevity launched a proprietary program called 100+ longevity program. The program claims to combine the most advanced technologies with a dedicated medical team to help people achieve their healthiest and longest life. [ 11 ]


In June 2022, Human Longevity announced an  initial public offering  through a  SPAC , with a valuation of $1 billion. [ 12 ]


The  International Congress of  Genetics  (ICG) is a five yearly conference for geneticists. The first ICG was held in 1898. Since 1973 It has been organized by the International Genetics Federation (IGF). The aim of the congress is to reflect on progress made in  genetics , to celebrate the best of contemporary research and to anticipate future developments in the discipline. It is one of the most important genetics meetings, presenting all subfields of the discipline. These subfields cover all present-day experiments using the powerful  genomic  technologies. Also, the benefits and wider implications of genetic research to societies at large are explored.


The Congress has been held in many major cities around the world, such as  London ,  New York City ,  Paris ,  Ithaca ,  Edinburgh ,  Stockholm ,  Bellagio ,  Montreal ,  The Hague ,  Tokyo ,  Berkeley ,  Moscow ,  New Delhi ,  Toronto ,  Birmingham ,  Beijing ,  Melbourne ,  Berlin  and  Singapore .


The last International Congress of Genetics took place in Singapore in 2013. In 2018, the 22nd International Congress of Genetics will be held in  Foz de Iguaçu , Brazil. [ 1 ]


The International Genetics Federation (IGF) is an association of national genetics societies interested in promoting the advancement of the science of genetics. The IGF is governed by a representative council that meets every five years at the International Congress of Genetics (ICG) to choose the venue of the next Congress, to elect IGF officers and members of the Executive Board, and to set dues, make or affirm policy decisions, and amend the IGF Constitution as needed.


The  HGSA Oration  was established in 1990 in order to honour a pre-eminent member of the  Human Genetics Society of Australasia (HGSA) . It is a lecture given by a senior member of the HGSA who has contributed significantly to the profession over many years. The oration is given annually at the  Annual Scientific Meeting of the HGSA . [ 1 ]


Metabarcoding   is the  barcoding  of  DNA / RNA  (or  eDNA / eRNA ) in a manner that allows for the simultaneous identification of many  taxa  within the same sample. The main difference between barcoding and metabarcoding is that metabarcoding does not focus on one specific organism, but instead aims to determine  species composition  within a sample.


A barcode consists of a short variable  gene  region (for example, see  different markers/barcodes ) which is useful for taxonomic assignment flanked by highly conserved gene regions which can be used for  primer  design. [ 1 ]  This idea of general barcoding originated in 2003 from researchers at the  University of Guelph . [ 2 ]


The metabarcoding procedure, like general barcoding, proceeds in order through stages of  DNA extraction ,  PCR amplification ,  sequencing  and  data analysis . Different genes are used depending if the aim is to barcode single species or metabarcoding several species. In the latter case, a more universal gene is used. Metabarcoding does not use single species DNA/RNA as a starting point, but DNA/RNA from several different organisms derived from one environmental or bulk sample.


Environmental DNA  or eDNA describes the genetic material present in environmental samples such as sediment, water, and air, including whole cells,  extracellular DNA  and potentially whole organisms. [ 3 ] [ 4 ]  eDNA can be captured from environmental samples and  preserved ,  extracted ,  amplified ,  sequenced , and categorized based on its sequence. [ 5 ]  From this information, detection and classification of species is possible. eDNA may come from skin, mucous, saliva, sperm, secretions, eggs, feces, urine, blood, roots, leaves, fruit, pollen, and rotting bodies of larger organisms, while  microorganisms  may be obtained in their entirety. [ 6 ] [ 7 ] [ 4 ]  eDNA production is dependent on  biomass , age and feeding activity of the organism as well as physiology, life history, and space use. [ 4 ] [ 8 ] [ 9 ] [ 10 ]


By 2019 methods in eDNA research had been expanded to be able to assess whole communities from a single sample. This process involves metabarcoding, which can be precisely defined as the use of general or universal  polymerase chain reaction  (PCR)  primers  on mixed DNA samples from any origin followed by  high-throughput   next-generation sequencing  (NGS) to determine the  species composition  of the sample. This method has been common in microbiology for years, but, as of 2020, it is only just finding its footing in the assessment of macroorganisms. [ 11 ] [ 12 ] [ 13 ]  Ecosystem-wide applications of eDNA metabarcoding have the potential to not only describe communities and biodiversity, but also to detect interactions and functional ecology over large spatial scales, though it may be limited by false readings due to contamination or other errors. [ 7 ] [ 14 ] [ 12 ] [ 9 ]  Altogether, eDNA metabarcoding increases speed, accuracy, and identification over traditional barcoding and decreases cost, but needs to be standardized and unified, integrating taxonomy and molecular methods for full ecological study. [ 11 ] [ 15 ] [ 16 ] [ 17 ] [ 9 ] [ 10 ]


eDNA metabarcoding has applications to diversity monitoring across all habitats and taxonomic groups, ancient ecosystem reconstruction, plant-pollinator interactions, diet analysis, invasive species detection, pollution responses, and air quality monitoring. eDNA metabarcoding is a unique method still in development and will likely remain in flux for some time as technology advances and procedures become standardized. However, as metabarcoding is optimized and its use becomes more widespread, it is likely to become an essential tool for ecological monitoring and global conservation study. [ 10 ]


Since the inception of high‐throughput sequencing ( HTS ), [ 19 ]  the use of metabarcoding as a biodiversity detection tool has drawn immense interest. [ 12 ] [ 20 ]  However, there has yet to be clarity regarding what source material is used to conduct metabarcoding analyses (e.g., environmental DNA versus  community DNA ). Without clarity between these two source materials, differences in sampling, as well as differences in laboratory procedures, can impact subsequent bioinformatics pipelines used for data processing, and complicate the interpretation of spatial and temporal biodiversity patterns. Here, we seek to clearly differentiate among the prevailing source materials used and their effect on downstream analysis and interpretation for environmental DNA metabarcoding of animals and plants compared to that of community DNA metabarcoding. [ 13 ]


With community DNA metabarcoding of animals and plants, the targeted groups are most often collected in bulk (e.g., soil, malaise trap or net), and individuals are removed from other sample debris and pooled together prior to bulk DNA extraction. [ 12 ]  In contrast, macro‐organism eDNA is isolated directly from an environmental material (e.g., soil or water) without prior segregation of individual organisms or plant material from the sample and implicitly assumes that the whole organism is not present in the sample. Of course, community DNA samples may contain DNA from parts of tissues, cells and organelles of other organisms (e.g., gut contents, cutaneous intracellular or extracellular DNA). Likewise, macro‐organism eDNA samples may inadvertently capture whole microscopic nontarget organisms (e.g., protists, bacteria). Thus, the distinction can at least partly break down in practice. [ 13 ]


Another important distinction between community DNA and macro‐organism eDNA is that sequences generated from community DNA metabarcoding can be taxonomically verified when the specimens are not destroyed in the extraction process. Here, sequences can then be generated from voucher specimens using Sanger sequencing. As the samples for eDNA metabarcoding lack whole organisms, no such in situ comparisons can be made. Taxonomic affinities can therefore only be established by directly comparing obtained sequences (or through bioinformatically generated operational taxonomic units (MOTUs)), to sequences that are taxonomically annotated such as NCBI's GenBank nucleotide database, [ 21 ]   BOLD , [ 22 ]  or to self‐generated reference databases from Sanger‐sequenced DNA. [ 23 ] [ 24 ] [ 25 ]  (The molecular  operational taxonomic unit  (MOTU) is a group identified through use of cluster algorithms and a predefined percentage sequence similarity, for example, 97%)). [ 26 ] [ 13 ]  Then, to at least partially corroborate the resulting list of taxa, comparisons are made with conventional physical, acoustic or visual‐based survey methods conducted at the same time or compared with historical records from surveys for a location (see Table 1). [ 13 ]


The difference in source material between community DNA and eDNA therefore has distinct ramifications for interpreting the scale of inference for time and space about the biodiversity detected. From community DNA, it is clear that the individual species were found in that time and place, but for eDNA, the organism that produced the DNA may be upstream from the sampled location, [ 27 ]  or the DNA may have been transported in the faeces of a more mobile predatory species (e.g., birds depositing fish eDNA, [ 28 ]  or was previously present, but no longer active in the community and detection is from DNA that was shed years to decades before. [ 29 ]  The latter means that the scale of inference both in space and in time must be considered carefully when inferring the presence for the species in the community based on eDNA. [ 13 ]


There are six stages or steps in DNA barcoding and metabarcoding. The DNA barcoding of animals (and specifically of  bats ) is used as an example in the diagram at the right and in the discussion immediately below.


First, suitable DNA barcoding regions are chosen to answer some specific research question. The most commonly used DNA barcode region for animals is a segment about 600  base pairs  long of the  mitochondrial gene   cytochrome oxidase I  (CO1). [ 2 ]  This locus provides large sequence variation between species yet relatively small amount of variation within species. [ 31 ]  Other commonly used barcode regions used for species identification of animals are  ribosomal DNA  (rDNA) regions such as  16S ,  18S  and  12S  and mitochondrial regions such as  cytochrome B . [ 32 ] [ 33 ] [ 34 ] [ 35 ]  These markers have advantages and disadvantages and are used for different purposes. [ 36 ] [ 37 ]  Longer barcode regions (at least 600 base pairs long) are often needed for accurate species delimitation, especially to differentiate close relatives. Identification of the producer of organism's remains such as faeces, hairs and saliva can be used as a  proxy measure  to verify absence/presence of a species in an ecosystem. The DNA in these remains is usually of low quality and quantity, and therefore, shorter barcodes of around 100 base pairs long are used in these cases. Similarly, DNA remains in dung are often degraded as well, so short barcodes are needed to identify prey consumed. [ 30 ]


Second, a reference database needs to be built of all DNA barcodes likely to occur in a study. Ideally, these barcodes need to be generated from vouchered specimens deposited in a publicly accessible place, such as for instance a natural history museum or another research institute. [ 30 ]  Building up such reference databases is currently being done all over the world. Partner organizations collaborate in international projects such as the  International Barcode of Life Project  (iBOL) and  Consortium for the Barcode of Life  (CBOL), aiming to construct a DNA barcode reference that will be the foundation for DNA‐based identification of the world's biome. Well‐known barcode repositories are  NCBI GenBank  and the  Barcode of Life Data System  (BOLD). [ 30 ]


Third, the cells containing the DNA of interest must be broken open to expose its DNA. This step,  DNA extractions  and  purifications , should be performed from the substrate under investigation. There are several procedures available for this. [ 38 ]  Specific techniques must be chosen to isolate DNA from substrates with partly degraded DNA, for example fossil samples, and samples containing inhibitors, such as blood, faeces and soil. Extractions in which DNA yield or quality is expected to be low should be carried out in an  ancient DNA  facility, together with established protocols to avoid contamination with modern DNA. [ 39 ] [ 40 ]  Experiments should always be performed in duplicate  [ 41 ]  and with positive controls included. [ 30 ]


Fourth,  amplicons  have to be generated from DNA extracted, either from a single specimen or from complex mixtures with  primers  based on DNA barcodes selected under step 1. To keep track of their origin, labelled  nucleotides  (molecular IDs or MID labels) need to be added in case of metabarcoding. These labels are needed later on in the analyses to trace reads from a bulk data set back to their origin. [ 30 ]


Fifth, the appropriate techniques should be chosen for  DNA sequencing . The classic  Sanger chain‐termination method  relies on the selective incorporation of chain‐elongating inhibitors of  DNA polymerase  during  DNA replication . These four bases are separated by size using  electrophoresis  and later identified by laser detection. The Sanger method is limited and can produce a single read at the same time and is therefore suitable to generate DNA barcodes from substrates that contain only a single species. [ 30 ]  Emerging technologies such as  nanopore sequencing  have resulted in the cost of DNA sequencing reducing from about USD 30,000 per megabyte in 2002 to about USD 0.60 in 2016. [ 43 ] [ 44 ]  Modern  next-generation sequencing  (NGS) technologies can handle thousands to millions reads in parallel and are therefore suitable for mass identification of a mix of different species present in a substrate, summarized as metabarcoding. [ 30 ]


Finally,  bioinformatic analyses  need to be carried out to  match DNA barcodes  obtained with  Barcode Index Numbers  (BINs) in reference libraries. [ 45 ]  Each BIN, or BIN cluster, can be identified to species level when it shows high (>97%) concordance with DNA barcodes linked to a species present in a reference library, or when taxonomic identification to the species level is still lacking, an  operational taxonomic unit  (OTU), which refers to a group of species (i.e. genus, family or higher taxonomic rank). [ 30 ]  (See  binning (metagenomics) ). The results of the bioinformatics pipeline must be pruned, for example by filtering out unreliable singletons, superfluous duplicates, low‐quality reads and/or  chimeric reads . This is generally done by carrying out serial  BLAST searches  in combination with automatic filtering and trimming scripts. [ 46 ]  Standardized thresholds are needed to discriminate between different species or a correct and a wrong identification. [ 30 ]


Despite the obvious power of the approach, eDNA metabarcoding is affected by precision and accuracy challenges distributed throughout the workflow in the field, in the laboratory and at the keyboard. [ 47 ]  As set out in the diagram at the right, following the initial study design (hypothesis/question, targeted taxonomic group etc) the current eDNA workflow consists of three components: field, laboratory and bioinformatics. [ 13 ]  The field component consists of sample collection (e.g., water, sediment, air) that is preserved or frozen prior to DNA extraction. The laboratory component has four basic steps: (i) DNA is concentrated (if not performed in the field) and purified, (ii)  PCR  is used to amplify a target gene or region, (iii) unique nucleotide sequences called "indexes" (also referred to as "barcodes") are incorporated using PCR or are  ligated  (bound) onto different PCR products, creating a "library" whereby multiple samples can be pooled together, and (iv) pooled libraries are then sequenced on a  high‐throughput machine . The final step after laboratory processing of samples is to computationally process the output files from the sequencer using a robust bioinformatics pipeline. [ 13 ]


The method requires each collected DNA to be archived with its corresponding "type specimen" (one for each taxon), in addition to the usual collection data. These types are stored in specific institutions (museums, molecular laboratories, universities, zoological gardens, botanical gardens, herbaria, etc.) one for each country, and in some cases, the same institution is assigned to contain the types of more than a country, in cases where some nations do not have the technology or financial resources to do so.


In this way, the creation of type specimens of genetic codes represents a methodology parallel to that carried out by traditional taxonomy.


In a first stage, the region of the DNA that would be used to make the barcode was defined. It had to be short and achieve a high percentage of unique sequences. For animals, algae and fungi, a portion of a mitochondrial gene which codes for subunit 1 of the cytochrome oxidase enzyme, CO1, has provided high percentages (95%), a region around 648 base pairs. [ 52 ]


In the case of plants, the use of CO1 has not been effective since they have low levels of variability in that region, in addition to the difficulties that are produced by the frequent effects of  polyploidy ,  introgression , and hybridization, so the chloroplast genome seems more suitable . [ 53 ] [ 54 ]


The diagram on the right shows a comparison of  pollination networks  based on DNA metabarcoding with more traditional networks based on direct observations of insect visits to plants. By detecting numerous additional hidden interactions, metabarcoding data largely alters the properties of the pollination networks compared to visit surveys. Molecular data shows that pollinators are much more  generalist  than expected from visit surveys. However, pollinator species were composed of relatively specialized individuals and formed functional groups highly specialized upon  floral morphs . [ 55 ]


As a consequence of the ongoing global changes, a dramatic and parallel worldwide  decline in pollinators  and animal-pollinated plant species has been observed. [ 56 ]  Understanding the responses of pollination networks to these declines is urgently required to diagnose the risks the ecosystems may incur as well as to design and evaluate the effectiveness of conservation actions. [ 57 ]  Early studies on animal pollination dealt with simplified systems, i.e. specific pairwise interactions or involved small subsets of plant-animal communities. However, the impacts of disturbances occur through highly complex interaction networks  [ 58 ]  and, nowadays, these complex systems are currently a major research focus. Assessing the true networks (determined by ecological process) from field surveys that are subject to sampling effects still provides challenges. [ 59 ] [ 55 ]


Recent research studies have clearly benefited from network concepts and tools to study the interaction patterns in large species assemblages. [ 60 ]  They showed that plant-pollinator networks were highly structured, deviating significantly from random associations. [ 61 ]  Commonly, networks have (1) a low connectance (the realized fraction of all potential links in the community) suggesting a low degree of generalization; (2) a high nestedness (the more-specialist organisms are more likely to interact with subsets of the species that more-generalist organisms interact with) the more specialist species interact only with proper subsets of those species interacting with the more generalist ones; [ 62 ]  (3) a cumulative distribution of connectivity (number of links per species, s) that follows a power or a truncated power law function  [ 63 ]  characterized by few supergeneralists with more links than expected by chance and many specialists; (4) a modular organization. A module is a group of plant and pollinator species that exhibits high levels of within-module connectivity, and that is poorly connected to species of other groups. [ 64 ] [ 55 ]


The low level of connectivity and the high proportion of specialists in pollination networks contrast with the view that generalization rather than specialization is the norm in networks. [ 65 ] [ 66 ]  Indeed, most plants species are visited by a diverse array of pollinators which exploit floral resources from a wide range of plant species. [ 67 ] [ 68 ]  A main cause evoked to explain this apparent contradiction is the incomplete sampling of interactions. [ 69 ]  Indeed, most network properties are highly sensitive to sampling intensity and network size. [ 61 ]  Network studies are basically phytocentric i.e. based on the observations of pollinator visits to flowers. This plant-centered approach suffers nevertheless from inherent limitations which may hamper the comprehension of mechanisms contributing to community assembly and biodiversity patterns. First, direct observations of pollinator visits to certain taxa such as orchids are often scarce  [ 70 ]  and rare interactions are very difficult to detect in field in general. [ 57 ] [ 58 ] [ 59 ] [ 60 ]  Pollinator and plant communities usually are composed of few abundant species and many rare species that are poorly recorded in visit surveys. [ 71 ] [ 72 ]  These rare species appear as specialists, whereas in fact they could be typical generalists. Because of the positive relationship between interaction frequency (f) and connectivity (s), undersampled interactions may lead to overestimating the degree of specialization in networks. [ 73 ]  Second, network analyses have mostly operated at species levels. Networks have very rarely been up scaled to the functional groups or down scaled to the individual-based networks, [ 74 ]  and most of them have been focused on one or two species only. The behavior of either individuals or colonies is commonly ignored, although it may influence the structure of the species networks. [ 74 ]  Species accounted as generalists in species networks could, therefore, entail cryptic specialized individuals or colonies. Third, flower visitors are by no means always effective pollinators as they may deposit no conspecific pollen and/or a lot of heterospecific pollen. [ 75 ] [ 76 ]  Animal-centered approaches based on the investigation of pollen loads on visitors and plant stigmas may be more efficient at revealing plant-pollinator interactions. [ 75 ] [ 76 ] [ 55 ]


Metabarcoding offers new opportunities for deciphering  trophic linkages  between predators and their prey within food webs. [ 78 ] [ 79 ]  Compared to traditional, time-consuming methods, such as  microscopic  or  serological analyses , the development of DNA metabarcoding allows the identification of prey species without prior knowledge of the predator’s prey range. [ 80 ]  In addition, metabarcoding can also be used to characterize a large number of species in a single  PCR  reaction, and to analyze several hundred samples simultaneously. [ 6 ]  Such an approach is increasingly used to explore the functional diversity and structure of food webs in agroecosystems. [ 79 ] [ 81 ] [ 82 ] [ 83 ] [ 84 ]  Like other molecular-based approaches, metabarcoding only gives qualitative results on the presence/absence of prey species in the gut or fecal samples. [ 85 ]  However, this knowledge of the identity of prey consumed by predators of the same species in a given environment enables a "pragmatic and useful surrogate for truly quantitative information. [ 86 ] [ 77 ]


In  food web ecology , "who eats whom" is a fundamental issue for gaining a better understanding of the complex  trophic interactions  existing between pests and their natural enemies within a given ecosystem. [ 36 ] [ 87 ]  The dietary analysis of arthropod and vertebrate predators allows the identification of key predators involved in the natural control of arthropod pests and gives insights into the breadth of their diet ( generalist vs. specialist ) and  intraguild predation . [ 77 ]


The diagram on the right summarises results from a 2020 study which used metabarcoding to untangle the functional diversity and structure of the food web associated with a couple of  millet  fields in Senegal. After assigning the identified OTUs as species, 27 arthropod prey taxa were identified from nine arthropod predators. The mean number of prey taxa detected per sample was the highest in  carabid beetles , ants and spiders, and the lowest in the remaining predators including  anthocorid  bugs,  pentatomid  bugs, and earwigs. Across predatory arthropods, a high diversity of arthropod preys was observed in spiders, carabid beetles, ants, and anthocorid bugs. In contrast, the diversity of prey species identified in earwigs and pentatomid bugs was relatively low.  Lepidoptera ,  Hemiptera ,  Diptera  and  Coleoptera  were the most common insect prey taxa detected from predatory arthropods. [ 77 ]


Conserving functional  biodiversity  and related  ecosystem services , especially by controlling pests using their natural enemies, offers new avenues to tackle challenges for the sustainable intensification of food production systems. [ 88 ] [ 89 ] [ 90 ]  Predation of  crop pests  by generalist predators, including arthropods and vertebrates, is a major component of natural  pest control . [ 91 ]  A particularly important trait of most generalist predators is that they can colonize crops early in the season by first feeding on alternative prey. [ 92 ] [ 93 ]  However, the breadth of the "generalist" diet entails some drawbacks for pest control, such as intra-guild predation. [ 91 ] [ 94 ] [ 95 ]  A tuned diagnosis of diet breadth in generalist predators, including predation of non-pest prey, is thus needed to better disentangle food webs (e.g., exploitation competition and apparent competition) and ultimately to identify key drivers of natural pest control in agroecosystems. However, the importance of generalist predators in the food web is generally difficult to assess, due to the ephemeral nature of individual predator–prey interactions. [ 94 ] [ 96 ]  The only conclusive evidence of predation results from direct observation of prey consumption, identification of prey residues within predators’ guts, [ 94 ]  and analyses of regurgitates [ 97 ]  or feces. [ 98 ] [ 77 ]


The spread of  non-indigenous species  (NIS) represents significant and increasing risks to ecosystems. [ 100 ]  In marine systems, NIS that survive the transport and adapt to new locations can have significant adverse effects on local biodiversity, including the displacement of native species, and shifts in biological communities and associated food webs. [ 101 ] [ 102 ]  Once NIS are established, they are extremely difficult and costly to eradicate, [ 103 ] [ 104 ]  and further regional spread may occur through natural dispersal or via anthropogenic transport pathways. [ 104 ] [ 105 ] [ 106 ]  While vessel hull fouling and ships’ ballast waters are well known as important anthropogenic pathways for the international spread of NIS, [ 100 ] [ 107 ] [ 108 ] [ 109 ]  comparatively little is known about the potential of regionally transiting vessels to contribute to the secondary spread of marine pests through bilge water translocation. [ 99 ]


Recent studies have revealed that the water and associated debris entrained in bilge spaces of small vessels (<20 m) can act as a vector for the spread of NIS at regional scales. [ 110 ] [ 111 ] [ 112 ] [ 113 ] [ 114 ] [ 115 ]  Bilge water is defined as any water that is retained on a vessel (other than ballast), and that is not deliberately pumped on board. It can accumulate on or below the vessel’s deck (e.g., under floor panels) through a variety of mechanisms, including wave actions, leaks, via the propeller stern glands, and through the loading of items such as diving, fishing, aquaculture or scientific equipment. [ 116 ]  Bilge water, therefore, may contain seawater as well as living organisms at various life stages, cell debris and contaminants (e.g., oil, dirt, detergent, etc.), all of which are usually discharged using automatic bilge pumps or are self-drained using duckbill valves. Bilge water pumped from small vessels (manually or automatically) is not usually treated prior to discharge to sea, contrasting with larger vessels that are required to separate oil and water using filtration systems, centrifugation, or carbon absorption. [ 116 ] [ 117 ]  If propagules are viable through this process, the discharge of bilge water may result in the spread of NIS. [ 99 ]


In 2017, Fletcher et al. used a combination of laboratory and field experiments to investigate the diversity, abundance, and survival of biological material contained in bilge water samples taken from small coastal vessels. [ 115 ]  Their laboratory experiment showed that  ascidian  colonies or fragments, and  bryozoan  larvae, can survive passage through an unfiltered pumping system largely unharmed. They also conducted the first morpho-molecular assessment (using eDNA metabarcoding) on the biosecurity risk posed by bilge water discharges from 30 small vessels (sailboats and motorboats) of various origins and sailing time. Using eDNA metabarcoding they characterised approximately three times more taxa than via traditional microscopic methods, including the detection of five species recognised as non-indigenous in the study region. [ 99 ]


To assist in understanding the risks associated with different NIS introduction vectors, traditional microscope biodiversity assessments are increasingly being complemented by eDNA metabarcoding. [ 118 ] [ 119 ] [ 120 ] [ 121 ]  This allows a wide range of diverse taxonomic assemblages, at many life stages to be identified. It can also enable the detection of NIS that may have been overlooked using traditional methods. Despite the great potential of eDNA metabarcoding tools for broad-scale taxonomic screening, [ 122 ] [ 123 ]  a key challenge for eDNA in the context of environmental monitoring of marine pests, and particularly when monitoring enclosed environments such as some bilge spaces or ballast tanks, is differentiating dead and viable organisms. [ 124 ]  Extracellular DNA can persist in dark/cold environments for extended periods of time (months to years, [ 125 ] [ 126 ]  thus many of the organisms detected using eDNA metabarcoding may have not been viable in the location of sample collection for days or weeks. In contrast, ribonucleic acid (RNA) deteriorates rapidly after cell death, likely providing a more accurate representation of viable communities. [ 127 ]  Recent metabarcoding studies have explored the use of co-extracted eDNA and eRNA molecules for monitoring benthic sediment samples around marine fish farms and oil drilling sites, [ 128 ] [ 129 ] [ 130 ] [ 131 ] [ 132 ]  and have collectively found slightly stronger correlations between biological and physico-chemical variables along impact gradients when using eRNA. From a marine biosecurity prospective, the detection of living NIS may represent a more serious and immediate threat than the detection of NIS based purely on a DNA signal.  Environmental RNA  may therefore offer a useful method for identifying living organisms in samples. [ 99 ]


The construction of the genetic barcode library was initially focused on fish  [ 133 ]  and birds, [ 134 ] [ 135 ] [ 136 ]  followed by butterflies and other invertebrates. [ 137 ]  In the case of birds, the DNA sample is usually obtained from the chest.


Researchers have already developed specific catalogs for large animal groups, such as bees, birds, mammals or fish. Another use is to analyze the complete zoocenosis of a given geographic area, such as the "Polar Life Bar Code" project that aims to collect the genetic traits of all organisms that live in polar regions; both poles of the Earth. Related to this form is the coding of all the ichthyofauna of a hydrographic basin, for example the one that began to develop in the Rio São Francisco, in the northeast of  Brazil . [ 138 ] [ 139 ]


The potential of the use of Barcodes is very wide, since the discovery of numerous cryptic species (it has already yielded numerous positive results), [ 140 ]  the use in the identification of species at any stage of their life, the secure identification in cases of protected species that are illegally trafficked, etc. [ 141 ] [ 142 ]


It has also been used as a non-invasive tool to determine the diet of wildlife species, such as wombats  [ 143 ]  and particularly critically endangered species, such as the  northern hairy-nosed wombat  ( Lasiorhinus krefftii ). [ 144 ]


DNA barcoding  has been proposed as a way to distinguish species suitable even for non-specialists to use. [ 145 ]


In general, the  shortcomings for DNA barcoding  are valid also for metabarcoding. One particular drawback for metabarcoding studies is that there is no consensus yet regarding the optimal experimental design and bioinformatics criteria to be applied in eDNA metabarcoding. [ 146 ]  However, there are current joined attempts, such as the COST network  DNAqua-Net  of the  European Cooperation in Science and Technology , [ 147 ]  to move forward by exchanging experience and knowledge to establish best-practice standards for biomonitoring. [ 148 ]


The so-called barcode is a region of mitochondrial DNA within the gene for  cytochrome c oxidase . A database,  Barcode of Life Data Systems  (BOLD), contains DNA barcode sequences from over 190,000 species. [ 149 ] [ 150 ]  However, scientists such as Rob DeSalle have expressed concern that classical taxonomy and DNA barcoding, which they consider a misnomer, need to be reconciled, as they delimit species differently. [ 151 ]  Genetic introgression mediated by endosymbionts and other vectors can further make barcodes ineffective in the identification of species. [ 152 ]


In  microbiology , genes can move freely even between distantly related bacteria, possibly extending to the whole bacterial domain. As a rule of thumb, microbiologists have assumed that kinds of  Bacteria  or  Archaea  with  16S ribosomal RNA  gene sequences more similar than 97% to each other need to be checked by  DNA-DNA hybridisation  to decide if they belong to the same species or not. [ 153 ]  This concept was narrowed in 2006 to a similarity of 98.7%. [ 154 ]


DNA-DNA hybridisation  is outdated, and results have sometimes led to misleading conclusions about species, as with the  pomarine  and  great skua . [ 155 ] [ 156 ]  Modern approaches compare sequence similarity using computational methods. [ 157 ]


The  National Centre for Plant Genetic Resources: Polish Genebank (NCPGR)  is a research unit in the Plant Breeding and Acclimatization Institute – National Research Institute. NCPGR is the coordinator and implementer of the National Crop Plant Genetic Resources Protection Programme. The Programme aims to protect the biodiversity of crop plants endangered by genetic erosion in Poland, and is funded by the Ministry of Agriculture. [ 1 ] [ 2 ]  The main tasks include collection of crop and wild plant populations and varieties threatened by genetic erosion, description and evaluation of collected materials, and preservation of their viability and genetic purity. The Programme is an implementation of provisions laid down in international treaties ratified by Poland:


NCPGR collects populations and cultivated varieties of crop and wild plants threatened with  genetic erosion . [ 3 ]  Collected materials are characterised, evaluated and documented.


Seed samples and clones are maintained in viable state and genetic purity. NCPGR exchanges samples with other institutions worldwide and provides initial plant materials for breeding and research programs. [ 4 ]


The Laboratory organizes collecting expeditions, during which plant genetic resources are obtained. Plants are collected from natural sites or obtained from farmers or on local markets. [ 5 ]  Collected material is reproduced and stored in a  gene bank . The Laboratory also carries out studies of variation and genetic structure of selected species and prepares initial materials of selected species for practical breeding. [ 6 ]


The Laboratory covers drawing up documentation of  genetic resources  of crop plants and exchanges information with other  genebanks . [ 7 ]  It also obtains seed samples of crop plants from national and international breeding centres. As for the long-term conservation, seed samples need to be prepared and have their viability tested before they are stored in temperature controlled chambers. NCPGR has three long-term storage chambers (-18 °C) and five medium-term storage chambers (0 °C).


The seeds exposed to long-term preservation are tested for viability and biochemical changes. Seed samples with low seed viability or with insufficient seed amount are forwarded for regeneration. The Laboratory provides seed samples for breeding, research and education. [ citation needed ]


The Laboratory searches for and collects local and foreign plant species with increased ability to adapt to extreme conditions of devastated environment. Another objective is regeneration and maintenance of viability of alternative plant collection for  reclamation  of devastated lands and lands periodically excluded from agricultural use. [ citation needed ]


Selected plant species are analyzed for having the ability to colonize different types of industrial and communal derivative areas. There is also description of morphological and other useful plant traits carried out when it comes to plants which have high biological yielding potential to serve as a source of energy and for other economic uses. [ citation needed ]


The Laboratory prepares initial material for the breeding programmes of alternative crops.


The key activity of Botanical Garden is evaluation and preservation of grass  genetic resources . The plants originated from various  climatic zones  are maintained in different, highly specialized collections. [ 8 ]  Natural resources of Botanical Garden include:


VIB  is a research institute located in  Flanders ,  Belgium . It was founded by the  Flemish government  in 1995, and became a full-fledged institute on 1 January 1996. The main objective of VIB is to strengthen the excellence of Flemish life sciences research and to turn the results into new economic growth. [ 1 ]  VIB spends almost 80% of its budget on  research  activities, while almost 12% is spent on  technology transfer  activities and stimulating the creation of new businesses, in addition VIB spends approximately 2% on socio-economic activities. VIB is member of EU-LIFE, an alliance of leading life sciences research centres in Europe. [ 2 ]


The institute is led by Christine Durinx and Jérôme Van Biervliet. [ 3 ] [ 4 ]   Ajit Shetty  is chairman of the board of directors. [ 5 ]


VIB's mission is  to conduct frontline biomolecular research in life sciences for the benefit of scientific progress and the benefit of society . The strategic goals of the VIB are:


The VIB scientist works on the normal and abnormal or pathological processes occurring in a cell, an organ and an organism (humans, plants, micro organisms). Instead of relocating scientists to a new campus, the VIB researchers work in research departments on six Flemish campuses:  Ghent University ,  KU Leuven ,  University of Antwerp ,  Vrije Universiteit Brussel ,  IMEC  and  Hasselt University .


VIB has established several core facilities focused on advanced technologies, which make high through-flow technologies available to academic and industrial researchers in Flanders.


VIB was involved in the creation of spin-offs from academic research groups, such as for  Ablynx ,  DevGen ,  CropDesign ,  ActoGeniX , Pronota (formerly Peakadilly), Agrosavfe, Multiplicom, Q-biologicals,  SoluCel , Aphea.Bio and Aelin Therapeutics. [ citation needed ] [ 6 ]


51°1′4.53″N   3°41′38.64″E  /  51.0179250°N 3.6940667°E  /  51.0179250; 3.6940667


The  Avery–MacLeod–McCarty experiment  was an experimental demonstration by  Oswald Avery ,  Colin MacLeod , and  Maclyn McCarty  that, in 1944, reported that  DNA  is the substance that causes  bacterial transformation , in an era when it had been widely believed that it was  proteins  that served the function of carrying genetic information (with the very word  protein  itself coined to indicate a belief that its function was  primary ).  It was the culmination of research in the 1930s and early 20th century at the  Rockefeller Institute for Medical Research  to purify and characterize the "transforming principle" responsible for the transformation phenomenon first described in  Griffith's experiment  of 1928: killed  Streptococcus pneumoniae  of the  virulent  strain type III-S, when injected along with living but non-virulent type II-R pneumococci, resulted in a deadly infection of type III-S pneumococci.  In their paper " Studies on the Chemical Nature of the Substance Inducing Transformation of Pneumococcal Types: Induction of Transformation by a Desoxyribonucleic Acid Fraction Isolated from Pneumococcus Type III ", published in the February 1944 issue of the  Journal of Experimental Medicine , Avery and his colleagues suggest that DNA, rather than protein as widely believed at the time, may be the hereditary material of bacteria, and could be analogous to  genes   and/or  viruses  in higher organisms. [ 1 ] [ 2 ]


With the development of  serological typing , medical researchers were able to sort bacteria into different  strains , or  types .  When a person or test animal (e.g., a  mouse ) is  inoculated  with a particular type, an  immune response  ensues, generating  antibodies  that react specifically with  antigens  on the bacteria.   Blood serum  containing the antibodies can then be extracted and applied to  cultured bacteria .  The antibodies will react with other bacteria of the same type as the original inoculation.  Fred Neufeld , a German bacteriologist, had discovered the pneumococcal types and serological typing; until  Frederick Griffith 's studies bacteriologists believed that the types were fixed and unchangeable from one generation to the next. [ 3 ]


Griffith's experiment , reported in 1928, [ 4 ]  identified that some "transforming principle" in pneumococcal bacteria could transform them from one type to another. Griffith, a British medical officer, had spent years applying serological typing to cases of  pneumonia , a frequently fatal disease in the early 20th century.  He found that multiple types—some virulent and some non-virulent—were often present over the course of a clinical case of pneumonia, and thought that one type might change into another (rather than simply multiple types being present all along).  In testing that possibility, he found that transformation could occur when dead bacteria of a virulent type and live bacteria of a non-virulent type were both injected in mice: the mice would develop a fatal infection (normally only caused by live bacteria of the virulent type) and die, and virulent bacteria could be isolated from such infected mice. [ 5 ]


The findings of Griffith's experiment were soon confirmed, first by  Fred Neufeld [ 6 ]  at the  Koch Institute  and by  Martin Henry Dawson  at the Rockefeller Institute. [ 7 ]  A series of Rockefeller Institute researchers continued to study transformation in the years that followed.  With Richard H. P. Sia, Dawson developed a method of transforming bacteria  in vitro  (rather than  in vivo  as Griffith had done). [ 8 ]  After Dawson's departure in 1930,  James Alloway  took up the attempt to extend Griffith's findings, resulting in the extraction of  aqueous solutions  of the transforming principle by 1933.  Colin MacLeod worked to purify such solutions from 1934 to 1937, and the work was continued in 1940 and completed by Maclyn McCarty. [ 9 ] [ 10 ]


Pneumococcus is characterized by  smooth  colonies which have a polysaccharide capsule that induces  antibody  formation; the different types are classified according to their immunological specificity. [ 1 ]


The purification procedure Avery  undertook consisted of first killing the bacteria with heat and  extracting  the  saline -soluble components.  Next, the protein was  precipitated  out using  chloroform  and the polysaccharide capsules were  hydrolyzed  with an  enzyme .  An immunological precipitation caused by type-specific antibodies was used to verify the complete destruction of the capsules.  Then, the active portion was precipitated out by alcohol  fractionation , resulting in fibrous strands that could be removed with a stirring rod. [ 1 ]


Chemical analysis showed that the proportions of carbon, hydrogen, nitrogen, and phosphorus in this active portion were consistent with the chemical composition of DNA.  To show that it was DNA rather than some small amount of  RNA ,  protein , or some other cell component that was responsible for transformation, Avery and his colleagues used a number of biochemical tests.  They found that  trypsin ,  chymotrypsin  and  ribonuclease  (enzymes that break apart proteins or RNA) did not affect it, but an enzyme preparation of "deoxyribonucleodepolymerase" (a crude preparation, obtainable from a number of animal sources, that could break down DNA) destroyed the extract's transforming power. [ 1 ]


Follow-up work in response to criticism and challenges included the purification and crystallization, by  Moses Kunitz  in 1948, of a DNA depolymerase ( deoxyribonuclease I ), and precise work by  Rollin Hotchkiss  showing that virtually all the detected nitrogen in the purified DNA came from  glycine , a breakdown product of the  nucleotide base   adenine , and that undetected protein contamination was at most 0.02% by Hotchkiss's estimation. [ 11 ] [ 12 ]


The experimental findings of the Avery–MacLeod–McCarty experiment were quickly confirmed, and extended to other hereditary characteristics besides polysaccharide capsules.  However, there was considerable reluctance to accept the conclusion that DNA was the genetic material.  According to  Phoebus Levene 's influential " tetranucleotide hypothesis ", DNA consisted of repeating units of the four nucleotide bases and had little biological specificity.  DNA was therefore thought to be the structural component of  chromosomes , whereas the genes were thought likely to be made of the protein component of chromosomes. [ 13 ] [ 14 ]   This line of thinking was reinforced by the 1935 crystallization of  tobacco mosaic virus  by  Wendell Stanley , [ 15 ]  and the parallels among viruses, genes, and enzymes; many biologists thought genes might be a sort of "super-enzyme", and viruses were shown according to Stanley to be proteins and to share the property of  autocatalysis  with many enzymes. [ 16 ]   Furthermore, few biologists thought that genetics could be applied to bacteria, since they lacked  chromosomes  and  sexual reproduction .  In particular, many of the geneticists known informally as the  phage group , which would become influential in the new discipline of  molecular biology  in the 1950s, were dismissive of DNA as the genetic material (and were inclined to avoid the "messy" biochemical approaches of Avery and his colleagues).  Some biologists, including fellow Rockefeller Institute Fellow  Alfred Mirsky , challenged Avery's finding that the transforming principle was pure DNA, suggesting that protein contaminants were instead responsible. [ 13 ] [ 14 ]   Although transformation occurred in some kinds of bacteria, it could not be replicated in other bacteria (nor in any higher organisms), and its significance seemed limited primarily to medicine. [ 13 ] [ 17 ]


Scientists looking back on the Avery–MacLeod–McCarty experiment have disagreed about just how influential it was in the 1940s and early 1950s.   Gunther Stent  suggested that it was largely ignored, and only celebrated afterwards—similarly to  Gregor Mendel 's work decades before the rise of  genetics .  Others, such as  Joshua Lederberg  and  Leslie C. Dunn , attest to its early significance and cite the experiment as the beginning of  molecular genetics . [ 18 ]


A few microbiologists and geneticists had taken an interest in the physical and chemical nature of genes before 1944, but the Avery–MacLeod–McCarty experiment brought renewed and wider interest in the subject.  While the original publication did not mention genetics specifically, Avery as well as many of the geneticists who read the paper were aware of the genetic implications—that Avery may have isolated the gene itself as pure DNA.  Biochemist  Erwin Chargaff , geneticist  H. J. Muller  and others praised the result as establishing the biological specificity of DNA and as having important implications for genetics if DNA played a similar role in higher organisms.  In 1945, the  Royal Society  awarded Avery the  Copley Medal , in part for his work on bacterial transformation. [ 19 ]


Between 1944 and 1954, the paper was cited at least 239 times (with citations spread evenly through those years), mostly in papers on microbiology, immunochemistry, and biochemistry.  In addition to the follow-up work by McCarty and others at the Rockefeller Institute in response to Mirsky's criticisms, the experiment spurred considerable work in microbiology, where it shed new light on the analogies between bacterial heredity and the genetics of sexually-reproducing organisms. [ 17 ]   French microbiologist  André Boivin  claimed to extend Avery's bacterial transformation findings to  Escherichia coli , [ 20 ]  although this could not be confirmed by other researchers. [ 17 ]   In 1946, however, Joshua Lederberg and  Edward Tatum  demonstrated  bacterial conjugation  in  E. coli  and showed that genetics could apply to bacteria, even if Avery's specific method of transformation was not general. [ 21 ]   Avery's work also motivated  Maurice Wilkins  to continue  X-ray crystallographic  studies of DNA, even as he faced pressure from funders to focus his research on whole cells, rather than biomolecules. [ 17 ]


Despite the significant number of citations to the paper and positive responses it received in the years following publication, Avery's work was largely neglected by much of the scientific community.  Although received positively by many scientists, the experiment did not seriously affect mainstream genetics research, in part because it made little difference for classical genetics experiments in which genes were defined by their behavior in breeding experiments rather than their chemical makeup.  H. J. Muller, while interested, was focused more on physical rather than chemical studies of the gene, as were most of the members of the  phage group .  Avery's work was also neglected by the  Nobel Foundation , which later expressed public regret for failing to award Avery a  Nobel Prize . [ 22 ]


By the time of the 1952  Hershey–Chase experiment , geneticists were more inclined to consider DNA as the genetic material, and  Alfred Hershey  was an influential member of the phage group. [ 23 ] [ 24 ]   Erwin Chargaff had shown that the base composition of DNA varies by species (contrary to the tetranucleotide hypothesis), [ 25 ]  and in 1952 Rollin Hotchkiss published his experimental evidence both confirming Chargaff's work and demonstrating the absence of protein in Avery's transforming principle. [ 26 ]  Furthermore, the field of  bacterial genetics  was quickly becoming established, and biologists were more inclined to think of heredity in the same terms for bacteria and higher organisms. [ 23 ] [ 24 ]   After Hershey and Chase used  radioactive isotopes  to show that it was primarily DNA, rather than protein, that entered bacteria upon infection with  bacteriophage , [ 27 ]  it was soon widely accepted that DNA was the material.  Despite the much less precise experimental results (they found a not-insignificant amount of protein entering the cells as well as DNA), the Hershey–Chase experiment was not subject to the same degree of challenge. Its influence was boosted by the growing network of the phage group and, the following year, by the publicity surrounding the DNA structure proposed by  Watson  and  Crick   (Watson was also a member of the phage group).  Only in retrospect, however, did either experiment definitively prove that DNA is the genetic material. [ 23 ] [ 24 ]


C7 protein  is an  engineered   zinc finger protein  based on the murine ZFP,  Zif268  and discovered by Wu  et al.  in 1994 (published in 1995). It shares the same  zinc finger  2 and zinc finger 3 of Zif268, but differs in the sequence of finger 1. It also shares the same DNA target, 5'-GCGTGGGCG-3'.


The shared sequences in  single letter amino acid codes  of fingers 2 and 3 are RSD-H-LTT and RAD-E-RKR (positions -1 through 6 in the alpha helix).


Zinc finger 1 has the sequence KSA-D-LKR which provides a 13-fold increase in affinity to the target sequence of the entire ZFP over that of Zif268.


It is used in zinc finger investigations in which the amino acid sequence of finger 2 is changed in order to determine the appropriate sequence to target a given three-nucleotide target site. A variation of C7,  C7.GAT  is preferred since it lacks the  aspartic acid  residue present in finger 3 of C7 and known to cause a phenomenon called 'target site overlap'. In this case the target site overlap is a result of the aspartic acid residue forming a hydrogen bond with the N4 of the cytosine (in the opposite strand) base-paired to the guanine in the finger 2 subsite. It can also form the same hydrogen bond with an adenine base paired to a thymine. This target site overlap would dictate that either a cytosine or adenine residue be present as the 3' nucleotide in the finger 2 subsite which is unacceptable when looking to target sequences containing another nucleotide at this position.


The  C7.GAT protein  is a  zinc finger protein  based on the  C7 protein  (itself based on the murine  Zif268 ). It features an alternative zinc finger 3  alpha helix  sequence, preventing the  target site overlap  caused by the  aspartic acid  residue of the finger 3 of C7. The sequence of this third finger is TSG-N-LVR according to the  single letter amino acid code . As the name suggest, the target site of finger 3 is altered to 5'-GAT-3', giving the overall protein a target of 5'-GCGTGGGAT-3'.


C7.GAT is used in studies investigating the effects of altering  zinc finger  alpha helix sequence on the target of the altered zinc finger, as well as the affinity and specificity of these proteins to their targets.


Cell ablation  (from Latin  Cellula  "small chamber" and  Ablatio  "removal"). Also known as  tissue  ablation ,  cell ablation is a biotechnological tool for studying cell lineage. The process consists of selectively destroying or removing cells in an organism.


For example, a laser beam or controllable gene promoter for a toxin gene can be used to destroy a selected amount of cells. [ 1 ]


Cell ablation can also be used as a tool to produce  transgenic  organisms lacking a cell type, and as a cure for certain diseases such as  cancer . [ 2 ]


The term is not to be confused with  genetic ablation : a method of modifying DNA in order to disrupt the production of a specific gene. [ 3 ]


There are multiple different techniques of cell ablation ( laser  ablation, thermal ablation, etc...). Many of which are being utilized in biomedical sciences to study cell functions [ 4 ]


Laser was used during the early stages of cell ablation, utilizing the flexibility, precision and heat generated from concentrated light waves (laser). Laser ablation was extremely effective in destroying cells  in vitro , as intensity of light was easily manipulated to injure cells. [ 4 ]  Laser ablation is not recommended for  in vivo , as the potential damage from laser was too high. High cost of operation, and extensive operating time narrowed its accessibility for cell studies. [ 4 ]


Recently, laser ablation was applied with  ICP-MS  to develop LA-ICP-MS analysis technique. In which utilizes the precision and ability to change laser intensity to determine elemental distribution of biological samples. [ 5 ]


Thermal ablation is widely used in biomedical science as a form of heat therapy for cancer. Extreme heat are applied to tumors, damaging the tissues to eliminate the tumors from eligible patients. [ 2 ]  Similarly, cryoablation can also be used as a potential treatment. Sudden drop to freezing temperature can cause crystal formation, puncturing membranes and causing cell death.


Radiofrequency ablation (RFA) utilizes high frequency sound waves to produce heat as a mean to destroy cells. RFA is frequently used as a minimally-evasive treatment for small tumors. [ 6 ]


Chimeric  nucleases are an example of  engineered proteins  which must comprise a  DNA-binding domain  to give sequence specificity and a  nuclease  domain for  DNA  cleavage.


DNA-binding domains including the  basic helix-loop-helix ,  zinc finger ,  helix-turn-helix  and  leucine zipper  motifs have been used in construction of sequence-specific nucleases. Of these, zinc fingers have been suggested the most important due to their modularity, allowing construction of a tailor-made DNA-binding domain. [ 1 ]


The nuclease domain is responsible for physical cleavage of DNA strands and may introduce either single stranded or double-stranded breaks.  FokI  is an example of a sequence-specific  endonuclease  whose non-specific nuclease domain introduces double stranded breaks and has been used in a variety of experiments including identification of high- and low-affinity binding sites of transcription factors  in vitro , to study recruitment of factors to promoter sites  in vivo  using  protein position identification with a nuclease tail  assay and to study proteins specific to interaction with DNA in the  Z-DNA  conformation (Durai et al., 2005 and references therein).


This  hydrolase  article is a  stub . You can help Wikipedia by  expanding it .


The  Crick, Brenner et al. experiment  (1961) was a  scientific experiment  performed by  Francis Crick ,  Sydney Brenner ,  Leslie Barnett  and R.J. Watts-Tobin.
It was a key experiment in the development of what is now known as molecular biology and led to a publication entitled "The General Nature of the Genetic Code for Proteins" and according to the historian of Science Horace Judson is "regarded...as a classic of intellectual clarity, precision and rigour". [ 1 ]  This study demonstrated that the  genetic code  is made up of a series of three base pair  codons  which code for individual  amino acids . The experiment also elucidated the nature of  gene expression  and  frame-shift mutations .


In the experiment,  proflavin -induced  mutations  of the  T4   bacteriophage  gene, rIIB, were isolated. Proflavin causes mutations by inserting itself between  DNA bases , typically resulting in insertion or deletion of a single base pair. [ 2 ]


Through the use of proflavin, the experimenters were able to insert or delete base pairs into their sequence of interest. When nucleotides were inserted or deleted, the gene would often be nonfunctional. However, if three base pairs were added or deleted, the gene would remain functional. [ 3 ]  This proved that the genetic code uses a  codon  of three nucleotide bases that corresponds to an  amino acid . The mutants produced by Crick and Brenner that could not produce functional rIIB protein were the results of  frameshift mutations , where the triplet code was disrupted.


Brenner and Crick et al. were also able to correct their frameshift mutations through the use of proflavin. If they had a nonfunctional gene due to a deleted base pair, by inserting a  base pair  into the general area of the deleted one, they were able to rescue the function of the gene. This is because the bases were shifted back into the correct  reading frame . [ 3 ]


In addition to discovering the triplet nature of the codon, this experiment confirmed the non-overlapping structure of the genetic code, identified the presence of "nonsense coding", and revealed the high degree of degeneracy in codon specification. [ 4 ]


This demonstration of the triplet nature of the genetic code, although carried out with bacteriophage, later proved to be universally applicable to all forms of life. [ 5 ]


Decades after its execution, the experiment remains a beacon of inspiration for scientists with its eloquent argumentation and straightforward yet impactful experiments, revealing fundamental insights into living organism organization. [ 6 ]  The results of this experiment inspired many to begin decoding the triplet code discovered by Brenner and Crick et al. Once this paper was published in 1961, researchers knew that there are 64 possible triplet codons, since there are four nitrogenous bases (4 x 4 x 4 = 64). Today, scientists have decoded what all 64 codons encode for, and the assignments have proven to be nearly universal. [ 5 ]


Genetic ablation  occurs when a gene is deemed “null” through the homologous genetic recombination of a gene. It is utilized in the selective suppression of a specific cell line or cell type. This genetic engineering technique does not limit growth suppression to just the activity of an individual gene. [ 1 ]  Specific cell ablation enables the examination of the  in vivo  activity of cells. An example of this method in action can be seen through the production of a knockout mouse. This is accomplished through the administration of one or more transgenes into a fertilized mouse oocyte’s pronucleus. Afterwards, it is reimplanted into a host mother, who then births a transgenic mouse. The transgenic mouse carries one copy of the transgene3 out of several hundred. From these mice, a homozygous colony can be created through breeding. [ 2 ]


In 1990, the  gene knockout  technique was just developing. There was a lack of information on the initial events that occur throughout the development of the vertebrate  embryo . In order to form a better understanding, the instructions for making an entire set of DNA in a person or organism need be dissected, and the genes involved with this process need to be determined. Instructions for embryonic development may have some correlation to the lack of space shown by many genes in their expression patterns. A technique used to evaluate specific gene function is through the inactivation or removal of that gene. By eradicating a specific gene, its role in development of the embryonic expression pattern may be able to be observed.  [ 3 ]


The ability to selectively remove cells by  ablation  is monumental in the study of the development of eukaryotic biology, contributing greatly to the study of the origin, fate, or function of the cells. Genetic ablation occurs through the delivery of a toxin or death-inducing gene that is directed by a cell-specific  enhancer (genetics) , or by utilizing the  GAL4/UAS system . Due to the array of known enhancers, toxins and death genes are able to be attached to nearly any cell selected, which permits cell-type-specificity. Through genetic ablation, the effects of removing every cell of a specific kind inside of an embryo are able to be observed; additionally, the entire population is able to be studied instead of just the individuals. [ 4 ]


Cell-type-specificity is a significant advantage of genetic ablation. The numerous enhancers that exist allow this specificity because toxins and death genes are able to target essentially any cell of choice. This cell specificity ablates all selected cell types in all sections of the embryo. This is an advantage because the number of analogous cells eliminated within a tissue has an influence on the phenotypic effects of ablation. Furthermore, because genetic ablation only requires organizing a genetic cross, it is simple technically, which allows a simultaneous examination of substantially sized populations of individuals. A larger number of samples helps authenticate the results, by providing more data to conclude from. Also, in certain cases, ablation is cell-autonomous, which eradicates any fear of compromising neighboring cells. This is seen in  ricin -A and  diphtheria -A chains as well as the death-inducing genes. [ 4 ]


There are also disadvantages associated with the genetic approaches to ablation. There is irregularity seen in expression that is driven by enhancers. These irregularities may be observed through a lack of restriction by a selected enhancer to a selected cell type or through a lack of inclusion for all cells of a certain kind in an embryo. Furthermore, the expressing cells can be killed by low levels of expression. A lack of choice of timing can prove to be a disadvantage as well. This is possible if the expression of the effector gene is GAL4- or enhancer-dependent. It is important to confirm that the toxin-encoding gene is only expressed during relevant developmental stages and in that specific cell in the embryo. This can be avoided by using mosaic expression. [ 4 ]


Temporal control of gene expression and ablation can be attributed to evolving  transgenic  and gene-therapy technologies. These technologies are enhanced by an understanding of the mechanisms that affect tissue-specific gene transcription. Genetic ablation allows for genes to be removed by compounds that are introduced into the organism of interest. [ 5 ]


Genetic ablation technology may be able to produce mice with mutations in just about every gene present in their germ line. Although this technique is not perfected, it contains the ability to target questions surrounding the molecular and cellular biology of embryonic growth. Furthermore, it may assist in the creation of animals to serve as guides showing the effects on human diseases including  demyelination ,  dwarfism , and  immunodeficiencies . [ 6 ]


Genetic ablation is a remarkable component in the study of cell lineages in mammals. This known quality encourages further study in dissecting plant developmental processes. A comprehensive overview of the stages in development is observed through intentional cell death by the use of promotors specifically shown in various cell types, along with the ability to produce genetically-engineered plants. Because of the more specific technique used in the production of chimeric plants, coupled with  laser ablation , genetic ablation serves as a principal mechanism for understanding plant cell development.  [ 7 ]


The  Hershey–Chase experiments  were a series of  experiments  conducted in 1952 [ 1 ]  by  Alfred Hershey  and  Martha Chase  that helped to confirm that  DNA  is  genetic material .


While DNA had been known to  biologists  since 1869, [ 2 ]  many scientists still assumed at the time that  proteins  carried the information for inheritance because DNA appeared to be an inert molecule, and, since it is located in the nucleus, its role was considered to be phosphorus storage. In their experiments, Hershey and Chase showed that when  bacteriophages , which are composed of DNA and protein, infect bacteria, their DNA enters the host bacterial cell, but most of their protein does not. Hershey and Chase and subsequent discoveries all served to prove that DNA is the hereditary material.


Hershey shared the 1969  Nobel Prize in Physiology or Medicine  with  Max Delbrück  and  Salvador Luria  for their "discoveries concerning the genetic structure of viruses". [ 3 ]


In the early twentieth century, biologists thought that proteins carried genetic information. This was based on the belief that proteins were more complex than DNA.  Phoebus Levene's  influential "tetranucleotide hypothesis", which incorrectly proposed that DNA was a repeating set of identical  nucleotides , supported this conclusion. The results of the  Avery–MacLeod–McCarty experiment , published in 1944, suggested that DNA was the genetic material, but there was still some hesitation within the general scientific community to accept this, which set the stage for the Hershey–Chase experiment. [ 4 ]


Hershey and Chase, along with others who had done related experiments, confirmed that DNA was the biomolecule that carried genetic information. Before that,  Oswald Avery ,  Colin MacLeod , and  Maclyn McCarty  had shown that DNA led to the transformation of one strain of  Streptococcus pneumoniae  to another. The results of these experiments provided evidence that DNA was the biomolecule that carried genetic information. [ 5 ]


Hershey and Chase needed to be able to examine different parts of the phages they were studying separately, so they needed to distinguish the phage subsections.  Viruses were known to be composed of a protein shell and DNA, so they chose to uniquely label each with a different elemental  isotope .  This allowed each to be observed and analyzed separately.  Since phosphorus is contained in DNA but not amino acids,  radioactive  phosphorus-32 was used to label the DNA contained in the  T2 phage .  Radioactive sulfur-35 was used to label the protein sections of the T2 phage, because sulfur is contained in protein but not DNA. [ 6 ]


Hershey and Chase inserted the radioactive elements in the bacteriophages by adding the isotopes to separate media within which bacteria were allowed to grow for 4 hours before bacteriophage introduction. When the bacteriophages infected the bacteria, the  progeny  contained the radioactive isotopes in their structures. This procedure was performed once for the sulfur-labeled phages and once for phosphorus-labeled phages.
The labeled progeny were then allowed to infect unlabeled bacteria. The phage coats remained on the outside of the bacteria, while genetic material entered. Disruption of phage from the bacteria by  agitation  in a blender followed by  centrifugation  allowed for the separation of the phage coats from the bacteria. These bacteria were  lysed  to release phage progeny. The progeny of the phages that were labeled with radioactive phosphorus remained labeled, whereas the progeny of the phages labeled with radioactive sulfur were unlabeled. Thus, the Hershey–Chase experiment helped to confirm that DNA, not protein, is the genetic material. [ 6 ]


Hershey and Chase showed that the introduction of  deoxyribonuclease  (referred to as  DNase ), an  enzyme  that breaks down DNA, into a solution containing the labeled bacteriophages did not introduce any  32 P into the solution.  This demonstrated that the phage is resistant to the enzyme while intact. Additionally, they were able to  plasmolyze  the bacteriophages so that they went into osmotic shock, which effectively created a solution containing most of the  32 P and a heavier solution containing structures called "ghosts" that contained the  35 S and the protein coat of the virus.  It was found that these "ghosts" could  adsorb  to bacteria that were susceptible to T2, although they contained no DNA and were simply the remains of the original viral capsule.  They concluded that the protein protected the DNA from DNase, but that once the two were separated and the phage was inactivated, the DNase could hydrolyze the phage DNA. [ 1 ]


Hershey and Chase were also able to prove that the DNA from the phage is inserted into the bacteria shortly after the virus attaches to its host. Using a high-speed blender they were able to force the bacteriophages from the bacterial cells after  adsorption . The lack of  32 P-labeled DNA remaining in the solution after the bacteriophages had been allowed to adsorb to the bacteria showed that the phage DNA was transferred into the bacterial cell.  The presence of almost all the radioactive  35 S in the solution showed that the protein coat that protects the DNA before adsorption stayed outside the cell. [ 1 ]


Hershey and Chase concluded that DNA, not protein, was the genetic material.  They determined that a protective protein coat was formed around the bacteriophage, but that the internal DNA is what conferred its ability to produce progeny inside a bacterium.  They showed that, in growth, protein has no function, while DNA has some function.  They determined this from the amount of radioactive material remaining outside of the cell.  Only 20% of the  32 P remained outside the cell, demonstrating that it was incorporated with DNA in the cell's genetic material.  All of the  35 S in the protein coats remained outside the cell, showing it was not incorporated into the cell, and that protein is not the genetic material. [ citation needed ]


Hershey and Chase's experiment concluded that little sulfur-containing material entered the bacterial cell.  However no specific conclusions can be made regarding whether material that is sulfur-free enters the bacterial cell after phage adsorption. Further research was necessary to conclude that it was solely bacteriophages' DNA that entered the cell and not a combination of protein and DNA where the protein did not contain any sulfur.


Hershey and Chase concluded that protein was not likely to be the hereditary genetic material.  However, they did not make any conclusions regarding the specific function of DNA as hereditary material, and only said that it must have some undefined role. [ 1 ] [ 7 ]