2.1. Comparison of Cp Genomes and Identification of Hypervariable Loci

All cp genome sequences in plants of the Zanthoxylum genus with complete genome sequence information were downloaded from GenBank (Table 1). The sequences were aligned using the Clustal W algorithm from MEGA7.0 [21] and CLC viewer 8.0 software (CLC bio, Aarhus, Denmark). The gene distribution patterns and similarities in the Zanthoxylum cp genomes were compared and visualized using mVISTA software (http://genome.lbl.gov/vista/mvista/submit.shtml) in Shuffle-LAGAN mode with the annotated Z. piperitum {"type":"entrez-nucleotide","attrs":{"text":"KT153018","term_id":"918056404","term_text":"KT153018"}}KT153018 cp genome as a reference. The variability of the aligned genomes was evaluated using the sliding window method with DnaSP ver. 5.0. software [22]. The window size was set to 600 base pairs (bp), the typical length of DNA markers. The step size was set to 300bp for relatively accurate positioning of hypervariable InDels. Only regions with a nucleotide diversity (Pi) value of >0.025 were considered. Hypervariable sites and genetic distance in the cp genome were calculated using MEGA 7.0. The InDel events were checked manually based on the aligned sequence matrix.

Chloroplast (cp) genomes of the four Zanthoxylum species used in this study.