DNA-SEQ of synthetic library for quantifying protein production: Regular condition, replicate #3
The synthetic E. coli library comprising 244,000 variants of a fluorescent reporter was grown in rich MOPS medium and binned into 16 populations of increasing fluorescence using FACS. The variable region of the reporter was PCR amplified from DNA extracted from each binned population. Primers used for generating the sequencing libraries contained handles for sequencing as well as custom sequencing barcodes for multiplexing. Libraries from all fluorescence bins were pooled to ratios reflecting their prevalence in the synthetic library and sequenced together (150PE). Resulting reads were demultiplexed through identification of the custom barcodes, which was then trimmed from the sequences along with the priming region. The fastq files contain these trimmed, demultiplexed reads. Reads with no assignable barcode were thrown away. No other data processing was performed.