PacBio amplicon re-sequencing of 62 P. tricornutum genomic loci

Re-sequencing of 62 endogenous P. tricornutum loci selected in a genome-wide analysis of haplotype diversity. The goal was to determine the number of haplotypes per locus and appearance of new haplotypes over time. The length of sequenced loci was 2kb (+/- 5%). Loci were amplified by emulsion PCR on the same culture harvested in two time points: five loci were amplified one month (T1) and all loci were amplified 6 months (T6) after the start of the culture from single cell. Plasmids containing cloned GFP or YFP were amplified separately as a control for random errors. Control reactions for artificial haplotypes detection consisted of mixed CFP with YFP or CFP with GFP. Amplicons were pooled together into two samples. Sample PacBio_AS_T1 contained five P. tricornutum endogenous amplicons from DNA harvested at T1 time point, GFP amplified separately and CFP+YFP amplified in one reaction. Sample PacBio_AS_T6 contained 63 P. tricornutum endogenous amplicons from DNA harvested at T6 time point, YFP amplified separately and CFP+GFP amplified in one reaction. Samples were mixed in 1:9 PacBio_AS_T1: PacBio_AS_T6 ratio before sequencing on one PacBio Sequel SMRT cell.