Losing DNA in purification?
917465
I always did the dephosphorylation simultaneously with the restriction. I used NEB restriction enzymes and shrimp alkaline phosphatase (SAP) from USB. As long as the total volume of enzymes as less than 10% of the reaction (to avoid too much glycerol in the mix) it worked fine.\n\nYou can heat inactivate XbaI. Also SAP is heat inactivated (though I haven't tried it). Check if CIP can be heat inactivated. If it can then probably you can get away with heat inactivation of the mix before using it for ligation. If however you have residual phosphatase activity then ligation will fail.\n\nIt looks like you have a medium or low copy number plasmid, but anyway since it is so big it sticks very well to the column.\nThe best choice would be to prepare lots of plasmid and afford a single purification step, just before doing the ligation.\nWhy don't you just use bigger culture volumes to get more plasmid? You could either do a midi or maxi prep, or use >1 mini-prep columns. \n\nI've been working with very low copy number plasmids so I know the problem. If you are cloning a PCR product I'd suggest that you first clone it in a simple high copy number vector like pBluescript, sequence it and then subclone it to your desired "problematic" vector. Otherwise you might get problems with the sequencing reaction-I've been there. If you are subcloning then OK.\n\nBy the way, the transformation efficiency for very big DNA fragments increases a lot if you add gyrase in the mix. I don't quite remember the citation (could be nucleic acids research) but I bet you can find it on Pubmed.
839482
A cell's DNA cannot be removed, but beta cerotene will remove free radicals, which are polyatomic ions that will react with things within the body which they shouldn't because they have a single electron which will readilt react with other ions and compounds to become stable.  DNA is in every cell and definatley cannot be removed.  Removal of parts of DNA result in mutations that are life threatening and con onlt happen at birth or due to radiation.
983150
Well, if you're using agarose you're looking at DNA.  THe higher concentrations of agarose separate smaller pieces of DNA more completely.\n\nLower concentrations generally cause smaller pieces to run together but allow for differentiation of bigger pieces.\n\nI tend to run 1% gels in TAE routinely, as measured by the DNA ladder we use (bands at 10k/8k/6k/5k/4k/2/k/1.5k/1.k/ 900/800/700/600/500/400/300/200/100)  that resolves everything pretty well except for maybe the 100-300 range, which can run off the gel if you're not careful.\n\nHowever, if I'm looking for size differences in smaller pieces, I'll run a 2% TAE gel (can tell 150 from 200 clearly, for example).  If I'm looking for size differences in bigger pieces, 6kb vs 6.2kb, I'll run a 0.7% TAE gel.\n\nThe molecular cloning manual has a chart for agarose concentrations and size range separations.
872183
You have 10 to 100 Trillion cells in your body.  Every cell has the same genetic code.  How would you change that code in so many cells?\n\nAdditionally, the genome is turning out to be much more complicated than was thought just a few years ago.  Much of a gene's 'performance' is determined by 'epigenetic' regulation of the gene's expression.  You can have 'normal' genes, but if they are not being expressed properly, that is the same as having defective genes.\n\nThis is a very, very complex issue.
304227
the nucleus??\ndna is stored??yes??
429879
Each half of the double-helix sort-of "unzips" during replication.  Since each of the bases is kind of like a jigsaw-puzzle piece, the two strands can then wind back up, if supplied with the appropriate free bases, to form two identical copies of the original DNA strand.\n\nNeat, huh?
335007
This is mostly true, but try it without RNAse, the yield is not that different.  It's for degrading residual RNAse I guess.  I think without RNAse the RNA and DNA bind together (not directly, but through huge protein/nucleic acid complexes), so the RNAse helps break this complex and release DNA.  Hope this makes sense.  It depends on the prufication procedure too, some just purify nucleic acid and rely on RNAse to get rid of RNA.  Some are specific for DNA. (ie qiagen).
1067028
Try a rough DNA isolation, some things u need are detergent, lemons, and a sample like onion.\n\nSimple diffusion experiments with onions, salt, water, etc..
43770
try these web sites or just type "purification protocol of mannose binding lectin" in to google
180398
It varies depending whether you want the final product as an active protein or denatured.\n\nFrom cells\n\nHomogenisation\nCentrifugation (assuming you know what fraction the protein will be in)\n\nYou could then possibley use centrifugation across a glucose gradient that would further separate proteins or more simpley...\n\nFind a pH or temperature where the protein is insoluble. (Extremes will denature the protein)\nor...\nUse a size exclusion coloumn to filter out larger smaller proteins (depending on the coloumns specifications and your needs, won't denature).\n\nFor further purification:\n\nUse isoelectric focusing across a gel.\nUse a affinity chromatography coloumn specific to your protein.\n\nHope that helps. It's from memory and should be ok, I study biochemistry so if I've things wrong then I should be quite worried!
730267
Mass-spectrometry (MS) is the only technique that gives true quantitative answers with extremely high certainty, but MS of DNA is uncommon and hard to do.  A more typical approach would be to "run a gel" separate the different fragments based on size: you have a porous matrix, you place the mixture of DNA pieces on one end, you apply a voltage.  Since the DNA is charged it migrates on the gel, but the bigger pieces are hindered by the matrix (the gel).  You then run a "standard" side-by-side with your unknown mixture--something with a mixture of a few fragments on known size.  Typically these "standard ladders" which are commercially available come in increments, e.g. 100 bp, 500 bp, 1000 bp, etc.  So you will be able to tell the size is between those fragment lengths (e.g. that a band from your unknown mixture is between 100 and 500 bp) with very high certainty, but you do not know the exact length with any certainty.\n\nAlternatively you can sequence DNA--this is very easy and cheap now-a-days, and has a very high-certainty.  So after doing a gel you can cut out the different bands and have them sequenced individually.  Both steps--the gel and the sequencing--are very routine, cheap, and reliable.
753317
Bored at work:\n\nYou could look for reverse transcriptase activity in infected cells.  This is a sure sign of RNA being turned into DNA.\n\nYou could purify virus from the aftermath of infected cells.  Then set up experiments to determine the nature of what you've isolated and what its properties are.
1098660
Your question isn't easy to read, but i think i understand it enough. There is one mole of 1kbp DNA strand with an EcoR1 restriction site in the middle. The amount of 5'ends produced when you addd EcoR1 to the DNA will be double because you will be effectively cutting the DNA in half. I hope this is what you were asking
652533
If by "sales" you meant "salts", then the answer is yes. DNA moves towards the cathode (+) due to its intrinsic negative charge at pH 7. If Na+ or K+ ions are at high concentration, then you would have the phosphate groups of DNA neutralized by those ions, and the overall charge of the molecule would be zero. Thus, no mobility through the gel.
729453
K.C.'s answer works if you have a selective media that will work. Also (or in addition to that) you may want to plate out colonies and then PCR out a unique sequence or two.\n\nThis wouldn't be definitive but if you design your primers right it may work, but also may be very time consuming.\n\nAlso you may want to try restriction digests to remap your gene. This would work well, if you have a good map.\n\nGood luck, what a bummer
901470
They're not available commercially. You have to purify them in a lab.
8687
mRNA  \nCapped messenger RNA is produced directly from your plasmid that contains a T7 or SP6 promoter. It is purified by our proprietary PUREmessenger™ technology. PUREmessenger™ is a chromatography that not only eliminates traces of DNA and proteins but also allows the selective recovery of the mRNA of the correct size. Shorter abortive transcripts as well as longer contaminating mRNA that are produced during in vitro transcription are eliminated. Capped mRNA will be purified by PUREmessenger™, quantified, re-suspended at 1mg/ml (except if otherwise specified) and aliquoted at 50 µl/aliquot (except if otherwise specified) in sterile conditions. It will be sent to you on cooling blocks together with its documentation: formaldehyde/agarose gel documentation picture. The capped mRNA that is sent to you ready-to-use (dissolved in water or the buffer of your choice) is 100% of your mRNA molecule of interest.\n\nCapped mRNA is produced in clean room facilities for research or clinical applications.\nYour benefit\nHighly purified, efficiently capped mRNA that has a superior efficacy compared to standard mRNA produced with in vitro transcription kits and simply purified by precipitation thus contaminated with alternative transcripts and traces of proteins and DNA fragments. The high quality of mRNA is reproduced from batch to batch. This guarantees the consistency of your results. Capped mRNA can be produced for research or for clinical utilisation: "documented" to be used as Active Pharmaceutical Ingredient or GMP-certified.