For the data obtained in the same lab you don't need to know a real length of fragments in nucleotides. You only need to know that this is one allele and that is an another. (But you must be sure that you sizing is reliable!) So, you can name the alleles as you want. You may use integer numbers in accordance of measured length. You may use number of repeats - take the control allele sequence from GenBank, see a number of flanking regions nucleotides and subtract it from measured length. In any case (without sequencing) your result will be conventional. So, you may simple name your alleles as 'a', 'b', 'c', d', etc. But there is no way to combine two labs data without analyzing of some reference samples. Or without sequencing made in both laboratories.
Hi Rayan, you need to sequence your PCR product. Only after you have the sequence you can see how many repetitions of your repeat are in each allele
The only way is to sequence the product (one allele if it is heterozygote). If there is this locus allele sequence in GenBank, you may use it, but for this case the size will be nominal. Because of the size, measured by comparing with size-standard may be not a result of simple nucleotides sum.
Strictly speaking, you need to sequence all your alleles to be sure. However, for some applications (i.e. population genetic diversity and differentiation statistics, coalescence based inference), you may use an estimate of number of repeats relative to the shortest allele. E.g. if you have alleles 120bp, 124bp and 126bp you may transform them into alleles 1 repeat, 3 repeats and 4 repeats (for a dinucleotide). Note than in some cases you might not be able to do that estimate (e.g. a dinucleotide with alleles 120bp, 121bp, 123bp...). Also, for other applications might not be appropriate.
Thank you all for your answers. Unfortunately I couldn't be able to sequence the PCR product right now. I'm trying to transform the PCR product size into number of repeats in order to compare our data with other populations. I found that the number of repeats differs in two populations using the same primers (20 repeats: 141bp, 20 repeats: 147bp). I have alleles 141bp, 143bp, 145bp...etc. So Miguel, would you please tell me in more details that how can I use an estimate of number of repeats relative to the shortest allele.
Rayan, can't you ask a reference sample from the other lab? If you do the PCR on a sample they already sized, and get your own alleles, then you solve your problem of comparison (if I correctly understood the problem). I.e. If they had an allele of 141bp, and for the same sample you get a size of 147bp, then you "just" have to add 6 to all your alleles to do the comparison.
If you want to treat your data and data obtained in another lab in one matrix - yes, you must unify alleles names ('sizes') at first - by analysing some reference samples. But what kind of samples you have? For some, labs are not glad to give their specimens to colleagues without collaboration agreement. But they may be gratefull to get yours.. And then they tell your an answer. :)
Good ideas. I'm working on human DNA. But the problem is that I'm studying in Sudan and no body else works in the same markers :(
Regarding my previous answer, I meant an estimate of *relative* number of repeats, not absolute number of repeats (for estimating genetic distances the difference in number of repeats is what counts). This estimate can only be done if all samples have been genotyped in the same lab or reference samples are used across labs.
I am not sure what do you mean with "the number of repeats differs in two populations using the same primers (20 repeats: 141bp, 20 repeats: 147bp)". How do you know the number of repeats for those alleles ? Were the two populations genotyped in two different labs ? Is there also an indel (of 6bp) in the amplified fragment ?
Also, you might also ask yourself if you really need the number of repeats. Diversity and differentiation statistics can be calculated from allele frequencies. Still you will have a problem if samples had been genotyped at different labs.
My samples have been analyzed in the same lab. But those of two populations were done at different labs. I don't know how did they count the number of repeats, I found them in the literature, and they didn't mention how. I need the number of repeats to be able to compare the allele frequencies of the Sudanese population with those of others. Because they all put the alleles in the form of number of repeats and not product size.
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