Is it possible to find out the integrity of DNA using real time PCR for comparison among two different species?
Hi Raksha,
you can easily run agarose gel to determine integrity of DNA. Also, if you try to see how they work in qPCR, you might check your result on agarose gel,. For your question I saw some pubs. online they have used mathematics values to compare DNAs integrity in different samples here is a link for one of them Article Incorporation of measurement of DNA integrity into qPCR assays
Good luck...
Integrity of which DNA? You have got the template and the PCR product. You won't be able to see the template-unless you put more than 100 ng in your reaction, which I doubt. As to the PCR product-I there assays using it for hybridization post RT PCR. So, if you did mean the template DNA than you have to use sufficient quantity to see it on the gel after the PCR.
the question is why do you want to check the integrity of DNA and as Leonid said which DNA? The DNA integrity is related to the template DNA and not the product. The information from the melt curve could itself give the information about the species variation, if Real-time is quantitative real time PCR, which I suppose it is.
Hi Raksha,
I am assuming you mean the integrity of the template DNA. Perhaps you are worried that your DNA sample might have suffered degradation.
If your template DNA is of good quality, then you will have a PCR product in your real-time reaction. If the DNA is degraded, you will not have amplified product. Be cautious, because if your amplification product is very short, it is possible to get amplification even from degraded DNA template.
However, I also would be very careful to check the PCR product by running the PCR product on a gel, staining the gel to visualize the DNA, extracting the expected product, and send for sequencing. This way you are sure that you are amplifying the correct material and not having mispriming or some other PCR "weirdness" taking place.
Good luck!
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