Would it be better to use cDNA or genomic DNA to run a conventional PCR on tissue from a goat?
it depends on the analysis which you are going to carry out further. If you are going for gene expression or protein expression studies then you must use cDNA as it has no non coding sequence.
If you want to genotype your goats you should use genomic DNA because several SNPS are in the noncoding region and couldn´t be found in cDNA.
sample source depend on what you are looking for ? if for example you need to study different expression profiles cDNA is better because it contains only all expressed DNA as mRNA but if you need to tact sequence or gene take genomic sample
Thanks. So I am gathering if I am doing sequencing, do genomic DNA, and if I am just looking for the presence of something, do cDNA, right.
To be even more specific, I want to look for a mutation in a specific gene, and plan to do genomic DNA and remove the band for sequencing. I also want to look for 2 viruses for possible etiologies of disease in the case, and I will then get cDNA for gel electrophoresis to see if the virus is present or not.
Isn't this correct?
Thank you all for your feedback. I have done many qRT-PCRs, but never conventional.
Using genomic DNA for sequencing in search of a mutation is correct. Ensure that your primers are designed so that they would not "miss" or mask the mutation (i.e. if you think the mutation is somewhere in the gene, ensure your primer sequences are designed to flank the entire gene). You do not have to isolate the band if your primers are designed around the entire gene itself. You can send the PCR product for sequencing (along with your non-mutated control), and compare the results.
As for the viruses, I would agree to use the cDNA as you have far higher copies of viral mRNA produced in the cells than the viral DNA. It would definitely make your life easier.
Hope this helps!
Hi Ebony! It depends on your ultimate aim. It is much simpler to prepare the genomic total DNA if you are the beginner with the amplifications.
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