Sequencing and Assembly of Chloroplast Genomes from Parasitic Plant Leaves

The experimental samples we collected were the fresh leaves of T. nigrans and S. parasitica, that parasitize on Platanus acerifolia and Ligustrum lucidum, respectively. The samples were collected at Sichuan University. We immediately froze those leaves in a liquid nitrogen tank and sent the frozen leaves to the Novogene Company for DNA extraction and genome sequencing. The modified CTAB method [27 ] was used to extract the total DNA of the leaf samples and Nanodrop was applied to detect DNA purity (OD 260/280 ratio). Sequencing was performed using the Illumina High-throughput Sequencing Platform (HiSeq/MiSeq). Finally, we obtained raw reads of 6.73 G and 6.85 G from T. nigrans and S. parasitica samples, respectively, and their GC contents were 42.3% and 41.6%, respectively.
We used the Trimmomatic v0.36 [28 (link)] to filter the raw reads and obtain high-quality clean reads. The BWA-MEM V0.7.12 [29 (link)] was used to compare the clean reads using T. chinensis as the reference chloroplast genome (Genbank: NC_036306.1). The read sequence was mapped to the corresponding reference genome. We used NOVOPlasty v2.6.3 [30 (link)] and Velvet v1.2.07 [31 (link)] to assemble and splice chloroplast genomes. We spliced contigs into scaffold sequences and then used them to assemble the chloroplast genome.