1. Look like the product does not indicate the DNA amount of each marker band. However, the instruction sheet does mention "Supplied at a concentration of 0.1µg/μl" (see attachment 1). However, I think this should be the concentration from all DNA fregments.

2. I suppose that you can cut out the 1 kb band and gel-purify it to measure its amount. But, it will be somehow inaccurate because DNA loss during purification.

3. If you really want to include marker DNA fragments with DNA amount, there are such products in the market. See the attached example (attachment 2) [http://www.enzolifesciences.com/ENZ-GEN103/ampigene-dna-ladder-250-10-000-bp/]. You can ask for a free sample from the company for testing. Most companies will be happy to provide one.

1. Look like the product does not indicate the DNA amount of each marker band. However, the instruction sheet does mention "Supplied at a concentration of 0.1µg/μl" (see attachment 1). However, I think this should be the concentration from all DNA fregments.

2. I suppose that you can cut out the 1 kb band and gel-purify it to measure its amount. But, it will be somehow inaccurate because DNA loss during purification.

3. If you really want to include marker DNA fragments with DNA amount, there are such products in the market. See the attached example (attachment 2) [http://www.enzolifesciences.com/ENZ-GEN103/ampigene-dna-ladder-250-10-000-bp/]. You can ask for a free sample from the company for testing. Most companies will be happy to provide one.

As it is indicated by @Yau that quantification with the gel is next to impossible directly. Furthermore, even with the quantification ladder, it would be practically not possible to estimate the band and to compare it with other band. The ladder is used for size, not for amount of DNA. How would you differentiate 20, 23 and 30 ng on gel. there are several factors related to see and interpret the gel. I guess, it useless to go for the quantification on gel idea, unless any external software based technique is used.

If you load 6 ul of 1 Kb ladder (Promega) => 600 ng/6 ul or 100 ng/ul ds DNA. If this is the concentration from all DNA fragments as Dr. Yuan-Yeu Yau said, then you can make a rough estimation.

You can make dilution (1/10, 1/100, 1/1000) either for your DNA unknown concentration or DNA known concentration (as a reference), afterwards run all of these samples in one gel include your 1 KB ladder.

For example, if you load 2 ul of undiluted DNA sample, then your sample concentration is : 100 ng/2 ul = 50 ng / ul. The easiest way and accurate one is using nanodrop for your DNA quantification.

Hope it’s useful.

Thanks, everyone for your insight and suggestions! I could buy a ladder that comes with quantified bands but I just wanted to salvage the one that I already have. My ballpark estimate for 1000 bp is 1/10 of total amount so ~10 ng/ul. I like the suggestion using nano drop after PCR clean-up. All I need is to get enough concentration for direct sequencing of 1.5kb PCR products.

This link is useful:

Article NanoDrop Microvolume Quantitation of Nucleic Acids

Regards

The simplest way to find out the concentration of the various bands would be to ask Promega. If you don't get an answer, you could scan the gel (or the image) and measure the area under each peak. Add the areas and normalise to the known concentration of total DNA, and you should get the concentration of DNA in each band. Scan the band(s) you want to measure and estimate concentration either by comparison to the most similar band or relative to the total area under peaks for the standard.

It's a bit roundabout and errors are sure to creep in, but it will give you a ballpark estimate.

Comparative gel electrophoresis is actually one of the most accurate methods of estimating DNA concentrations of very small amounts of DNA as it is much less affected by impurities than spectrophotometric methods.

Try ImageJ and get an estimate about the ratio proportion of your band of interest. Fermentas ladder generally have the quantity given for each, which I use for quantitation. For example, if you have 10 bands and your band density is 1/10th of total, then it is 60 ng (assuming you load 600 ng in 6 ul). But, be careful with the PCR band thickness, it should lie in the range of your ladder bands, othewise, it will be wrong estimation. Make dilution of your PCR and bring the intensity in the range of your ladder band intensity.

Many years ago, I had an information sheet from Promega that listed the supposed mass of each of these bands (I have been using this marker for over twenty years). Though I have searched for it, I can not find it (and the one copy taped to our gel camera was pilfered). I still have a couple of places to look, when I get a chance.

I will say that I remember the brightest bands (1000[?] and 3000) were ~100 ng each. I think one was ~96-98 ng and the other was ~103 ng. Lighter bands (most of the others) were in the 52-63 ng range with the lightest bands being ~28-32 ng (the 250/253 and 500 bands, I believe). I also believe there was one band in the 40s somewhere, but I do not remember which band it was. Of course, this was for a 5 ul load of the marker.

We used these weights by visual comparison to bands of interest in our gels and the weights usually proved fairly accurate when we checked our DNA with a spec. Now NanoDrop specs preclude such estimates.

Assuredly, I recall that only one band was a fraction more than 100 ng with another being a fraction less. Since you asked for a "rough" estimation, I hope this will be of some use to you.

I don't know if I will have a chance to dig into old notes more than 20 years old. However, now I am curious about this. If I find the info sheet, I will surely post it for you.

agree with Yuan-Yeu Yau

regards