基于转录组测序的青藏扁蓿豆EST-SSR标记开发与验证

Development and verification of EST-SSR markers in Medicago archiducis-nicolai by transcriptome sequencing

青藏扁蓿豆(Medicago archiducis-nicolai)广泛分布于青藏高原及其毗邻地区,对高海拔极端环境具有极强的适应性。本研究利用高通量转录组测序数据,对青藏扁蓿豆的EST-SSR标记进行了开发。结果发现,随机选取的477对引物中,350对可在青藏扁蓿豆中有效扩增,其中346对引物在扁蓿豆(M.ruthenica)中可实现跨物种扩增。利用其中64对独立遗传且符合哈迪–温伯格平衡的标记引物对青藏扁蓿豆和扁蓿豆野生群体的遗传多样性和群体遗传结构分析发现,两个物种存在明显的种间和种内的遗传分化,且提示种内群体间的遗传分化与地理隔离或环境选择有关。本研究所开发的EST-SSR标记为今后进一步评价青藏扁蓿豆和扁蓿豆的遗传多样性以及解释其适应极端环境的遗传机制提供了重要的研究工具。

Medicago archiducis-nicolai,which is endemic to the Qinghai-Tibet Plateau and its adjacent regions,is a species of Platycarpae in the genus Medicago.This species is characterized by its superior adaptability to extremely harsh environments.In this study,we developed EST-SSR markers of M.archiducis-nicolai using transcriptome sequencing.Polymerase chain reactions verified 350 of 477 randomly chosen EST-SSR primer sets in M.archiducis-nicolai.Of the verified primer sets,346 were successfully amplified in M.ruthenica,indicating high transferability of the EST-SSR primers to M.ruthenica.The genetic diversity and structure of M.archiducis-nicolai and M.ruthenica populations were characterized using 64 non-genetically linked EST-SSR loci that did not deviate from Hardy-Weinberg equilibrium.Results show significant inter-and intra-species genetic differentiation of the two species,implying geographic distance and environmental selection influenced intra-species genetic differentiation.The EST-SSR markers developed in this study provide useful tools for detailed characterization of the genetic diversity of M.archiducis-nicolai and M.ruthenica and reveal the mechanisms underlying their adaptation to harsh environments.