12 votes
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Are alleles equally sized?

In general, alleles don't have to be the same size. Two major examples which come to mind are the Huntingtin gene and FMR1. Huntingtin is the causative gene of Huntington's disease. In people with ...
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  • 1,534
11 votes
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DNA extraction from agarose gel

Yes, this is possible (and I have done it uncounted number of times) - the method is called "Freeze and squeeze". What you basically do is to run the gel, cut out the band of interest (be careful with ...
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10 votes
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What is the best file format to store gel images?

Always keep the raw data files! This is always a good idea for scientific data, and the only exception should be if the raw data is prohibitively large and it is not feasible to store it completely. ...
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6 votes
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How do you visualize RNA on a gel?

You have two possibilities: When you only need a quick check if your RNA is ok and you indeed only get one band, you can try a "quick and dirty" method. Heat the sample for 5 minutes at 65°C and then ...
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6 votes

What is possibly wrong with my gel electrophoresis when i don't see bands of DNA ladder?

As one attempts their daily Sisyphean challenge of uncovering a new fact there is one truism, or platitude, that informs all of our efforts at troubleshooting an unanticipated experimental outcome. ...
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  • 3,407
5 votes

Possible reasons for DNA getting stuck in well

There is one simple reason for that, your agarose gel is most likely too dense. Depending in the type of agarose, I would prepare a 0.5-0.6% gel at maximum. Synbio gives this list for "standard" ...
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5 votes

Why does hemoglobin electrophoresis in sickle cell trait gives two bands but not three?

This is a really interesting question and again reminds of how ignorant we are about things as subtle as this which otherwise should have been an obvious question. First of all, although there seems ...
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  • 3,636
5 votes
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How does SDS-PAGE separate based on mass?

You are correct that molecule mobility depends on the mass-to-charge ratio, and this means that different sized molecules with the same $\frac{m}{q}$ will have the same acceleration. However, the ...
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  • 7,564
4 votes

Appropriate Buffer for electerophoresis of DNA & Protein TBE or TAE?

There are already many great answers to your question, however I thought I put my comments in form of an answer. The standard for DNA agarose gel is TAE and for the protein, it depends on the size of ...
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4 votes

Appropriate Buffer for electerophoresis of DNA & Protein TBE or TAE?

Grossly, it does not matter what buffer you use. It is the pH that matters. For DNA electrophoresis EDTA is added in order to chelate divalent cations that serve as cofactors for nucleases. Tris is ...
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4 votes
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Appropriate Buffer for electerophoresis of DNA & Protein TBE or TAE?

The question which buffer for DNA is better is quite old. Both have their pros and cons and I list a few of them: TBE is a better conductor and is thus less prone for overheating the gel Borate is a ...
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4 votes

Appropriate Buffer for electerophoresis of DNA & Protein TBE or TAE?

I have had good experience using a lithium boric acid buffer from Faster Better Media. I use it for RNA gels, but it's advertised for DNA gels. I don't think it can do protein, but I've never tried it....
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4 votes

How bad is ethidium bromide in your plasmid for downstream applications?

Purification and isolation of DNA bands by cutting them from agarose gel is commonplace (Lee et al., 2012). The purification step after excision of the band gets rid of most of the EtBr and other ...
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4 votes
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Why would I see no bands in DNA gel electrophoresis of red blood cells?

It is a trick question. Red Blood Cells (RBC) are anucleated in mammals. You are not going to be able to extract anything about chromosome 1 from RBC. By the way, if you want to know how they got ...
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4 votes
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Are longer and shorter DNA similarly charged?

Each phosphate group has a single negative charge. Not just the terminals. The reason gel electrophoresis works so well with DNA is that charge is linearly proportional to size. The longer the ...
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3 votes
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Long term storage of agarose-ethidium bromide gels that have already been cast

To increase storage life: after gel solidifies, dampen it with running buffer. Wrap the gel in polyvinyl chloride. Place in plastic container with a lid. Store in fridge in dark. It will last for a ...
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  • 3,675
3 votes
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Troubleshooting SDS-PAGE of trypsin-treated BSA

I'm going to treat this as a partial homework question but provide some guidance as to how you can potentially address your question and have solid theory to back it up. Chymotrypsin preferentially ...
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3 votes

Single stranded DNA in gel electrophoresis

As already mentioned formamide can be used. However formamide has to be incubated with the nucleic acid sample and heated. Urea can be included in the gel (8%). DNA can also be denatured using NaOH. ...
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3 votes
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Protocol to dilute DNA step ladder?

You'll probably have to titrate it down yourself. You might be able to estimate: The detection limit for ethidium bromide staining is about $1 ng$ per band. The insert says it's at "1 $\mu g/ \mu l$", ...
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3 votes
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Why does my gel have such poor resolution?

Although your tech is referring to your DNA samples, your ladder also indicates there is some room for improvement of your running technique. -20C for a week should not have impacted the quality of ...
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  • 344
3 votes
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What do we call this adjustable platform used to ensure that something is positioned strictly level in a lab?

I remember using those for agarose gels as an undergraduate, though using them for PAGE doesn't make a lot of sense unless your bench is seriously out of level. Anyways, they can be called levelling ...
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3 votes

Quantifying DNA in a band on gel electrophoresis

All three methods could be used to measure the amount of DNA. However in practice, method 2 (estimation by dye brightness) typically works best in a normal workflow. It really depends on what you plan ...
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  • 344
3 votes

Agarose gel Ladder smear

Are you sure that your TBE is at the right dilution? And that you're using TBE to make the gel and also as the buffer for running? That sort of wavy line looks like what you might get if you'd made ...
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3 votes

Why is gel used in electrophoresis?

In gel electrophoresis, the gel is the mechanism by which macromolecules of different sizes are separated. By loading the gel with amino acids (or proteins or DNA), you start all of the samples ...
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  • 111
3 votes
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What is meant by 4 –12% or 8% SDS-PAGE?

Do the percentage values refer to the percentage of acrylamide in the gel? Yes. The 8% gel is 8 g acrylamide per 100 mL. The “4-12%” gel is a gradient gel, which are useful for separating proteins ...
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3 votes
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Electrodes in non-conducting fluid? Are they neutral? Electrolysis, Electrophoresis

Electrodes are neutral, but they are held at different potential. This is similar to the current flow in a wire: although current flows, the wire remains neutral, i.e., no charge is accumulated (...
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2 votes

What causes skewed lanes in a DNA gel electrophoresis experiment?

@Chris's suggestion is very possible, high salt characteristically shows this towards the end of the gel. But there are additional suspects. This looks like it's just an agarose gel, Correct? I've ...
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  • 3,675
2 votes

Gel Electrophoresis - loading dye and more

The loading buffer you add to your samples for gel electrophoresis has a few different purposes, but the exact amount does not really matter. The purpose of the loading buffer is to make the sample ...
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2 votes
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A question on ImageQuantTL

I would definitely go for option A. If you are assuming that the staining in band 6 is all due to whatever you are measuring, there is no reason to think that the signal at the top of lane 1 is not ...
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