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11

Short answer: Yes Long answer: Depends what you are working with. DNA: If you are working with DNA, its pretty stable and you can usually get away with a 70% ethanol wash/autoclave (mainly to prevent contamination and obtain consistent results). EDIT: Read Chris's answer also below RNA: If you are working with RNA well.. whatever you did for DNA doesnt ...


11

Well, don't use M or B, those are already taken (C or A, and not A, respectively). You can see the full list here: http://www.dna.affrc.go.jp/misc/MPsrch/InfoIUPAC.html (The enWiki article on Nucleobases lists a few others but I would ignore those as 1. D is present in both and 2. they are rare and inapplicable) 5-methylcytosine isn't on there. If you ...


9

It may seem counterintuitive that deoxyribonucleic acid has nitrogenous bases. Nonetheless, nucleic acids (thus including RNA) were called that way because the phosphate backbone (linked by phosphodiester groups) is a derivative of phosphoric acid. Phosphoric acid Image Source Phosphodiester group Image Source Note that the only difference between ...


8

First of all, sterility is not necessary. It takes much more effort to reach this than just to wipe down everything with ethanol. What you need is a clean and controlled work environment (but this is something you need anyway to get reproducible results) and good and clean equipment. You will need more precautions for RNA work as RNA is more sensitive and ...


8

The pKas of (neutral) guanine and thymine are 9-10 (ref). At high pH (>~10), those bases will be deprotonated and exist as negatively charged conjugate bases. As the deprotonated species, part of the G/C and A/T hydrogen bonding networks are eliminated. In the figure below, green dotted lines represent the hydrogen bonds that explain the observed base ...


8

A list of dyes is available here and a list of dyes specific to nucleic acids is available here. I think you have two choices the SYBR Gold nucleic acid gel stain (S11494) which can be detected under UV light (not sure if it can be used with polyacrylamide gels). Your other option which can be used with polyacrylamide gels is the SYBR Green II RNA gel stain ...


7

I would think GelRed or GelGreen would be an option too. They claim to be more sensitive than EtBr and certainly less toxic (even moreso than SYBR Green). I haven't personally used them against such a small bp product though. GelRed has basically the same excitation/emission wavelengths as EtBr so no equipment change is needed. Product sheet Store link


6

I always did mine at -80 C, but I never compared the results to other protocols (I don't fix what's not broken). But, I was curious about the same thing, so I looked around. I found one paper discussing this: Paithankar and Prasad, Nuc Acids Res 19(6):1346 (1991) It shows that at low concentrations, EtOH at RT actually outperforms the precipitations at both ...


6

There is indeed a repulsive force between the phosphates in the DNA backbone. (The pK of the phosphate is very low so it will be ionised at all physiologically-relevant pH.) This is why DNA behaves like a stiff rod over short distances. However cations in solution will help to shield some of this repulsion. This is why DNA melting temperature decreases as ...


5

This is a question of chemical nomenclature and the principle source for this is the IUPAC (IUBMB in case of biological molecules; but not in this case). You can find all hetero-ring nomenclature references on the IUPAC web site: http://www.acdlabs.com/iupac/nomenclature/ http://www.acdlabs.com/iupac/nomenclature/79/r79_702.htm (I'm skipping the actual ...


5

Correct, all but one step in pyrimidine synthesis occurs in the cytosol, and purine synthesis occurs in the cytosol only. Nucleotides freely diffuse through nuclear pores but are actively transported through mitochondrial membranes via embedded membrane transport proteins (i.e. ATP-ADP translocators, GTP transporters, and pyrimidine nucleotide ...


4

RNAses are enzymes, and there are various ways to inactivate them. Unfortunately, RNAses are rather stable proteins and autoclaving doesn't completely kill their activity. The common methods to inactivate are unspecific methods that will destroy any enzymes, either through covalent modification or degradation. The most common methods for RNAse inactivation ...


3

As user137 said, the general base abstracts a proton from the 2'OH and subsequently the 2'O- renders a nucleophilic attack on the δ+ Phosphorous, leading to the hydrolysis of the phosphodiester bond. There can be slight variations in the mechanism and the intermediates; for details see this review. ...


3

B-form DNA is wrapped around histones in a left-handed manner resulting in a left-handed solenoidal superhelix (see that, that and this). The reason for this wrapping is that it reduces the helical tension. This post has more information about DNA helical tension. Also note that exceptions exist (i.e. right-ended direction) especially for histone at the ...


3

First, you need to know which genome sequence does the SNP file refer to. They must have mentioned the reference sequence that they used. As others mentioned the case of CT is heterozygosity. If you just want to mark the changes then discard the residue that is already present in the reference genome and use the other allele. However, you want to keep a ...


3

The only paper I found that examined the question is "Structural, Dynamical, and Electronic Transport Properties of Modified DNA Duplexes Containing Size-Expanded Nucleobases". They state The results confirm that the structural and flexibility properties of the canonical DNA are globally little affected by the presence of benzo-fused bases. The most ...


2

Just to fully document the IUPAC advice on this, here is the relevant section from the Nomenclature for Incompletely Specified Bases in Nucleic Acid Sequences as reproduced here in the link provided by @Hamish McWilliam. 5.2. Modified nucleotides In a number of organisms DNA and RNA are modified at certain positions. For instance, the DNA of ...


2

As Amory suggests the IUPAC "Recommendations on Organic & Biochemical Nomenclature, Symbols & Terminology etc." (http://www.chem.qmul.ac.uk/iupac/) are probably the best place to start. From a quick scan the obviously relevant documents would be: "Abbreviations and Symbols for Nucleic Acids, Polynucleotides and their Constituents" "Nomenclature for ...


2

If you're looking for an exact match, you don't really need a complex aligner. Perl regular expressions are pretty powerful at string transformations or conditional matching of substrings. For example, to find all matches of AASYWSRA in a nucleotide sequence $seq, you can do: @matches = $seq =~ m/AA[CG][CT][AT][CG][AG]A/g; The [] brackets are known as ...


2

If sufficient time has elapsed, there will be a large number of transversions present between the original base and the final state. The initial state will therefore not matter. Say, you flip (turn over, not toss) a coin at random intervals. For short time and few flips, the initial state will matter. But if you go on flipping it for millenia, the final ...


2

As a first year molecular biology student, I did a class project on this very question: trying to see if buffer temperature had an effect on yield during DNA precipitation. This is a far cry from peer reviewed research, but my finding was that there was no significant effect for a wide range of temperatures.


1

As to why they don't recognize RNA-DNA heteroduplexes (which are present during transcription, for example), I suspect that the methylation which protects bacterial genomic dsDNA (see the DNA modifying enzyme section of this Columbia University lecture for more info) also protects RNA-DNA hybrids, as the genomic DNA would still be methylated. Note that most ...


1

This paper found that these enzymes recognize RNA:DNA heteroduplexes. Such duplexes are unlikely to be encountered in vivo. They are present when DNA is primed for replication, but these duplexes are relatively short and thus are less likely to randomly contain a recognition sequence. Furthermore, if the recognition sequence is found in the primer, the gDNA ...



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