I am using a Promega SV genomic DNA extraction kit with a modified protocol for extracting Gram positive bacteria. See link for details: www.sjsu.edu/people/peter.nemes/courses/Chem131A/s2/Genomic-DNA-Purification-Protocol.doc
I am finding that when I use the eluted DNA in qPCR the reaction is being inhibited. I suspect that a contaminant is being eluted with the DNA, so I did a NaAc/EtOH precipitation and then repeated the qPCR, and the samples produced the expected response. I’m reluctant to routinely use the NaAC/EtOH going forward, because I will be quantifying the DNA in the reaction and this additional step will increase error in the results.
My plan going forward is to dilute my DNA 1:100 to reduce the contaminant, but my curious mind want to actually find a solution.
Has anyone else experienced this problem?
Does anyone have any suggestions potential sources of contaminants from spin column DNA extractions?
Thanks for your help!