Chloroplast alignment and phylogenetics
Quality filtered and trimmed reads were mapped to the Mentha
longifolia plastome (GenBank NC_032054, masking one of the inverted
repeats) using bowtie2 v.2.3.4.3 (Langmead & Salzberg, 2012; Langmead,
Wilks Antonescu, & Charles, 2019) with default settings. Mapped reads
were quality filtered using SAMtools v.1.9 (Danecek et al., 2021)
removing all reads mapping with a mapping quality score below 20 and PCR
duplicated reads were removed using PicardTools v.2.25.2
(https://github.com/broadinstitute/picard). For each sample a consensus
sequence was extracted using BCFtools v.1.9 (Danecek et al., 2021) and
all ambiguous bases were recorded as N. Plastomes were aligned in mafft
v.7.392 (Katoh & Standley, 2013) with default settings and a
bootstrapped maximum-likelihood phylogenetic tree was inferred in RaxML
v.8.2.11 (Stamatakis, 2014) with a GTR+G+I mutation model usingSalvia an outgroup (Li et al., 2016; NCBI Genebank accessions:
NC_050898, MN520020, MN520021, NC_038165, MG772529, MN520019,
NC_050897, MN520023, and NC_050900).