Development of 10 SSR markers for a Tibetan medicinal plant Lancea tibetica (Phrymaceae) based on RAD sequencing
We selected one individual from each of the populations and pooled them. Subsequently, the RAD library was constructed based on published methods (Barchi et al., 2011). The library was quantified with Qubit (Invitrogen, Eugene, OR, USA) and sequenced using the Illumina MiseqTM platform (Illumina, San Diego, CA, USA). Before doing any further analysis, quality control and filtering of raw data were performed to detect whether the raw reads were of high enough quality, following Zhang et al. (2014). After that, clean reads were clustered using cd-hit-est (Li and Godzik, 2006) and assembled de novo using Velvetopt (Zerbino & Birney, 2008). Sequencing produced 2,764,204,500 bp of clean reads after quality control from 2,800,948,250 bp of raw reads. We obtained 1,417,277 cluster tags, but only 222,628 cut cluster tags. We obtained 401,203 high-quality contigs, with an average size of 265 bp (N50 = 361) through de novo assembly.