I have been doing some qPCRs on bacterial genomic DNA. I am using 16S as my "housekeeping gene". My sample of interest, however, yield a lower CT value.
I am trying to figure out how to overcome this issue.
Should I redesign 16S primers?
Is there any other "housekeeping gene" for bacteria?
Should I send my samples to genome sequencing?
There is no issue. Your bacteria contain many molecules of 16S rRNA, what results in low Ct values. That's all. This is not a problem.
There may only be a problem with this when your treatment leads to considerable changes in the amount of protein biosynthesis for which the amount of ribosomes and hence 16S rRNA may be regulated. A regulated housekeeping gene is not valid reference gene. The expression level (-> Ct value) is irrelevant - as long as it is constant across the treatments/samples you like to compare. Note that any gene being constantly exprseed under all the conditions/samples you like to compare would be a valid reference gene (even if it might be not expressed under some other circumstances - so that it would not be a housekeeping gene).
You're using genomic DNA, and yet you've picked one of the few genes that has a high and potentially variable copy number as your reference.
Perhaps this was not the best choice?
This aside, what is you're actually trying to do? Usually the applications of qPCR to genomic DNA are...fairly limited: things like copy number, or genotyping, or maybe for estimating the fraction of a large population of cells that has a given genotype.
Having said all this, there's no a priori problem with Cq values varying between samples: the whole point of a reference is to account for variation between samples, so provided you know your reference is either stable (cDNA) or of known and essentially invariant copy number (gDNA), the variation you see reflects differences in starting material, allowing you to normalise for that accordingly.
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