GSM5646617: SC_low; Saccharomyces cerevisiae; OTHER
After FACS-sorting and overnight regrowth in the same media used for growth, sorted fractions were miniprepped. PCR was used to verify that at least 10 plasmids were extracted per sorted cell; minipreps were PCR-amplified, further amplified with the addition of bin-specific barcodes, pooled, and amplified briefly again. For each sample, >50ng purified PCR fragment was used for library preparation. These PCR products were treated with End Prep Enzyme Mix for end repairing, 5' Phosphorylation and dAtailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Adaptor-ligated DNA was then performed using DNA Clean Beads. A second PCR reaction was carried out with P5 and P7 primers carrying sequences which can anneal with flowcell to perform bridge PCR and index allowing for multiplexing. The final library product for sequencing were then purified by beads and was qualified.