Chapter 9 Libraries preparation results

9.1 Post-enrichment PCR1 quantification

After the enrichment and the purification of the first PCR (PCR1), we quantified double strand DNA in every plates in order to adjust the second PCR (PCR2), and more especially in order to increase the number of cycles in PCR2. We used both Quant-It and few samples on Qbit and transformed raw absorbance results into concentration with regressions.

9.2 Post-enrichment (PCR1 & PCR2) and amplification quantification

Library showing no band or light smir in the electrophoresis have been reamplified, resulting in amplified plates A1 and A2. We used the Quant-It to dose all samples (original libraries and reamplified libraries).

9.3 Amplification result

We used electrophoresis and noticed if re-amplified sampled had a band (B), a light smir (L) or nothing (A). Samples without anything (A) had their library remade.

Figure 9.1: Electrophoresis Amplified

9.4 Library & extraction repetition result

69 libraries were still not ok after re-amplification. Those library have been rebuilt from source DNA. Among them, 43 were still not good at the electrophoresis and have bee re-extracted. Electrophoresis of re-extracted samples show a high heterogeneity of size, and around 8 of them seem to have not worked. All new samples need to be quantified..