Performed qPCR on the same exact samples previously sent to Alice Nguyen at the Marine Science Institute for digital droplet PCR (ddPCR) to compare our results to hers. These samples were previously quantified as part of the Ab Endo Project (Ab Endo 2011 Water Filter DNA samples). Here’s the list of samples that were sent to (and back from) Alice:
- CI SRI CP 1A
- CARMEL +500M 2
- CI SRI CP 2B
- CI SRI CP 2A
- CI SRI CP 1B
- CARMEL +500M 1
Master mix calcs are here: 20150415 – qPCR WS Ab Endo vs ddPCR
Standard curve was p16RK7 (from 20120730).
Cycling params, plate layout, etc. can be found in the qPCR Report (see Results below).
Table – Summary of qPCRs and ddPCR data
|Sample Number||Sample Name||Initial qPCR Copies||ddPCR Copies||Today qPCR Copies|
|AB01||CI SRI CP 1A||0||3.8||0|
|AB02||CARMEL +500M 2||1.57||8.7||1.3|
|AB03||CI SRI CP 2B||147||104||59.2|
|AB04||CI SRI CP 2A||0||175||83.1|
|AB05||CI SRI CP 1B||74.8||2.8||0|
|AB06||CARMEL +500M 1||1.08||4.9||0|
The table certainly shows some inconsistency, both between ddPCR and qPCR. Additionally, there is also inconsistency between the initial qPCRs and today’s qPCRs. Due to the limited sample volumes remaining for these sub-samples that were sent to Alice and then returned to us, I will repeat the qPCR using the stock DNA and see how that compares.