Hello,
I have been working on using qPCR as part of a project to determine relative telomere length in birds. However, my r2 and efficiencies have been inconsistent, sometimes returning appropriate results (r2 ~ .9, Eff ~ 100%), but other times repeating the same procedures returns significantly worse results.
I've attempted several times to eliminate pipetting error as a source of this inconsistency, but was wondering if there are alternative explanations besides this.
Pranay Agarwal
Hi. Have you tried running the qPCRs using a freshly prepared set of standards? Repeated freeze/thaw cycles may affect the quality of standard DNA preps as well. I personally use a set of standards for running ~3 qPCRs and then prepare a new one.
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