SSR是经典的分子标记之一，被广泛用于系统发育分析、分子辅助育种、遗传图谱构建、新品种鉴定等分析。高通量、高精准的SSR分型是进行李论研究和实践运用的基础，但是由于受到传统的分型方法（凝胶用，毛细管电泳、一代测序等）的制约，SSR分型一直处于低通量的状态，且在样品间比较时只能依赖于电泳图谱的相似程度进行，在耗费大量人力物力的情况下依然不能获得准确的分型结果。我院李论研究员基于扩增子测序技术开发了一套全新的SSR分型方法，可以实现在单碱基水平分型，且一次实验可以实现对100个品种的几千个SSR位点同时进行准确分型，大大降低了分型成本和耗费的时间，同时提高了分型准确性；样品间的比较可以直接基于碱基序列进行，实现了任意两个品种间的比较，避免了反复电泳的麻烦。目前该研究成果发表在《核酸研究》（IF：10.16）上。
Abstract
Accurate and efficient genotyping of simple sequence repeats (SSRs) constitutes the basis of SSRs as an effective genetic marker with various applications. However, the existing methods for SSR genotyping suffer from low sensitivity, low accuracy, low efficiency and high cost. In order to fully exploit the potential of SSRs as genetic marker, we developed a novel method for SSR genotyping, named as AmpSeq-SSR, which combines multiplexing polymerase chain reaction (PCR), targeted deep sequencing and comprehensive analysis. AmpSeq-SSR is able to genotype potentially more than a million SSRs at once using the current sequencing techniques. In the current study, we simultaneously genotyped 3105 SSRs in eight rice varieties, which were further validated experimentally. The results showed that the accuracies of AmpSeq-SSR were nearly 100 and 94% with a single base resolution for homozygous and heterozygous samples, respectively. To demonstrate the power of AmpSeq-SSR, we adopted it in two applications. The first was to construct discriminative fingerprints of the rice varieties using 3105 SSRs, which offer much greater discriminative power than the 48 SSRs commonly used for rice. The second was to map Xa21, a gene that confers persistent resistance to rice bacterial blight. We demonstrated that genome-scale fingerprints of an organism can be efficiently constructed and candidate genes, such as Xa21 in rice, can be accurately and efficiently mapped using an innovative strategy consisting of multiplexing PCR, targeted sequencing and computational analysis. While the work we present focused on rice, AmpSeq-SSR can be readily extended to animals and micro-organisms.
全文链接：https://academic.oup.com/nar/article/45/10/e88/2981921
