De novo Chloroplast Genome Assembly

Assembly of complete cp genome and nrDNA sequences was performed by de novo assembly of the low coverage whole genome sequence (WGS) via a bioinformatics pipeline (http://phyzen.com). Briefly, trimmed reads with Phred scores of 20 or less were prepared from the total pair-end (PE) raw reads using the CLC-quality trim tool and then were assembled by a CLC genome assembler (ver. 4.06 beta, CLC Inc, Rarhus, Denmark) with parameters of minimum 200 to 600 bp autonomously controlled overlap size. The principal contigs representing the cp genome were retrieved from the total contigs using MUMmer [18 (link)] with the cp genome sequence of Panax ginseng cv. ChP (KM088019) as reference sequence. The representative cp contigs were arranged in order based on the previously reported cp genome sequence and connected into a single draft sequence by joining overlapping terminal sequences. Assembly errors were identified in the initial assembly contigs and manually corrected by mapping of raw reads to assembled sequences. Error correction was validated by nucleotide sequencing after PCR amplification.