Application of high-throughput amplicon sequencing-based SSR genotyping in genetic background screening

文献类型： 外文期刊

第一作者： Li, Tiantian

作者： Li, Tiantian;Fang, Zhiwei;Peng, Hai;Zhou, Junfei;Zhu, Wenhui;Li, Lun;Chen, Lihong;Li, Lili;Liu, Zhihao;Zhang, Weixiong;Lu, Long;Gao, Lifen;Liu, Pengcheng;Wang, Yanyan;Zhai, Wenxue;Zhang, Quanfang;Lu, Long

作者机构：

关键词： SSR-based genetic background screening; Xa21; Marker-assisted backcrossing; Transgenesis; CRISPR; Cas9

期刊名称：BMC GENOMICS （ 影响因子：3.969； 五年影响因子：4.478 ）

ISSN： 1471-2164

年卷期： 2019 年 20 卷

页码：

收录情况： SCI

摘要： BackgroundHost genetic backgrounds affect gene functions. The genetic backgrounds of genetically engineered organisms must be identified to confirm their genetic backgrounds identity with those of recipients. Marker-assisted backcrossing (MAB), transgenesis and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9) editing are three commonly used genetic engineering techniques. However, methods for genetic background screening between genetically engineered organisms and corresponding recipients suffer from low efficiency, low accuracy or high cost.ResultsHere, we improved our previously reported AmpSeq-SSR method, an amplicon sequencing-based simple sequence repeat (SSR) genotyping method, by selecting SSR loci with high polymorphism among varieties. Ultimately, a set of 396 SSRs was generated and applied to evaluate the genetic backgrounds identity between rice lines developed through MAB, transgenesis, and CRISPR/Cas9 editing and the respective recipient rice. We discovered that the percentage of different SSRs between the MAB-developed rice line and its recipient was as high as 23.5%. In contrast, only 0.8% of SSRs were different between the CRISPR/Cas9-system-mediated rice line and its recipient, while no SSRs showed different genotypes between the transgenic rice line and its recipient. Furthermore, most differential SSRs induced by MAB technology were located in non-coding regions (62.9%), followed by untranslated regions (21.0%) and coding regions (16.1%). Trinucleotide repeats were the most prevalent type of altered SSR. Most importantly, all altered SSRs located in coding regions were trinucleotide repeats.ConclusionsThis method is not only useful for the background evaluation of genetic resources but also expands our understanding of the unintended effects of different genetic engineering techniques. While the work we present focused on rice, this method can be readily extended to other organisms.

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[1]Application of high-throughput amplicon sequencing-based SSR genotyping in genetic background screening. Li, Tiantian,Fang, Zhiwei,Peng, Hai,Zhou, Junfei,Zhu, Wenhui,Li, Lun,Chen, Lihong,Li, Lili,Liu, Zhihao,Zhang, Weixiong,Lu, Long,Gao, Lifen,Liu, Pengcheng,Wang, Yanyan,Zhai, Wenxue,Zhang, Quanfang,Lu, Long. 2019

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