Data from: A set of plastid loci for use in multiplex fragment length genotyping for intraspecific variation in Pinus (Pinaceae)

Premise of the study: Recently released Pinus plastome sequences support characterization of 15 plastid Simple Sequence Repeat (ptSSR) loci originally published for P. contorta and P. thunbergii. This allows selection of loci for single-tube PCR multiplexed genotyping in any subsection of the genus. Methods: Unique placement of primers and primer conservation across the genus were investigated, and a set of six loci were selected for single-tube multiplexing. We compare interspecific variation between ptSSRs and nucleotide sequences of ycf1 then test intraspecific variation for ptSSRs using 911 samples in the P. ponderosa species complex. Results: The ptSSR loci contain mononucleotide and complex repeats with additional length variation in flanking regions. They are not located in hypervariable regions and most primers are conserved across the genus. A single PCR per sample multiplexed for six loci yielded 45 alleles in 911 samples. Discussion: The protocol allows efficient genotyping of many samples. The ptSSR loci are too variable for Pinus phylogenies but are useful for the study of genetic structure within and among populations. The multiplex method could easily be extended to other plant groups by choosing primers for ptSSR loci in a plastome alignment for the target group.

研究背景：近期发布的松属（Pinus）质体基因组（plastome）序列，为解析此前针对扭叶松（P. contorta）和黑松（P. thunbergii）发表的15个质体简单序列重复（plastid Simple Sequence Repeat，ptSSR）位点提供了支撑。这使得我们能够为该属任何亚组的单管聚合酶链式反应（Polymerase Chain Reaction，PCR）多重基因分型筛选合适的位点。
方法：我们对引物的独特定位及该属内的引物保守性进行了研究，并筛选出6个位点用于单管多重扩增。我们对比了ptSSR位点的种间变异与ycf1基因的核苷酸序列，随后利用黄松物种复合体（P. ponderosa species complex）的911份样本，对ptSSR位点的种内变异进行了检测。
结果：ptSSR位点包含单核苷酸重复与复合重复序列，其侧翼区域存在额外的长度变异。此类位点并不位于高变区域内，且大多数引物在松属内具有保守性。针对911份样本进行单管6重PCR扩增，共获得45个等位基因。
讨论：本实验方案可实现大量样本的高效基因分型。ptSSR位点的变异程度过高，不适用于松属的系统发育研究，但可用于种群内部及种群间的遗传结构分析。通过针对目标类群的质体基因组比对筛选ptSSR位点引物，该多重扩增方法可轻松推广至其他植物类群。