目的 基于赤霉素（gibberellin A₃，GA₃）处理下的南方红豆杉Taxus chinensis var. mairei转录组数据鉴定和分析SSR位点，开发可用于南方红豆杉种质资源鉴定和遗传多样性分析的SSR标记。方法 使用MISA软件对南方红豆杉转录组数据进行SSR分析，搜索SSR位点。可搜索单核苷酸、二核苷酸、三核苷酸、四核苷酸、五核苷酸和六核苷酸，最小重复数分别设置为10、6、5、5、5、5次。采用Primer 3（2.3.5版，默认参数）进行SSR引物设计。通过毛细管荧光电泳检测，最终选定多态性好，扩增成功率高的SSR引物用于后续数据分析；通过MEGA进行UPGMA聚类分析，构建聚类树。结果 转录组测序共获得202 361条Unigene，采用MISA软件搜索出12 008个SSR位点，分布在10 894条Unigenes上，发生频率为5.38%，平均距离为1/7.64 kb。单核苷酸重复类型最丰富，占总SSR位点的51.74%，其次为三核苷酸（20.92%）和二核苷酸重复类型（20.88%）。A/T、AG/CT是优势重复基序，分别占总SSR重复类型的49.60%和10.93%。随机抽选96对SSR引物，以4个地区12份南方红豆杉种质进行引物有效性和多态性验证，筛选出13对具多态性的SSR引物，多态性引物比率为13.54%。13对引物在东北红豆杉Taxus cuspidata扩增效率高达100%，但在曼地亚红豆杉Taxus × media扩增效率仅为23.08%。聚类分析表明太行山地区南方红豆杉聚为一类，浙江、福建和湖南等南方来源的南方红豆杉聚为一类，后三者彼此之间遗传距离更近。结论 南方红豆杉转录组测序产生的Unigene信息可以用来开发SSR分子标记，开发的13个标记将有助于南方红豆杉物种遗传多样性、分子育种、资源保护等方面的研究。

Objective To identify and analyze SSR loci based on the transcriptome data of Taxus chinensis var. mairei, Lemee et levl S. Y. Hu et Liu treated with gibberellin A3 (GA₃), and to develop SSR markers that can be used for the identification of the germplasm resources and genetic diversity analysis of T. chinensis var. mairei. Method Perform SSR analysis on transcriptome data of southern Chinese yew using MISA software and search for SSR loci. Single nucleotide, dinucleotide, trinucleotide, tetranucleotide, pentanucleotide, and hexanucleotide can be searched, with the minimum number of repetitions set to 10, 6, 5, 5, 5, and 5, respectively. Primer 3 (version 2.3.5, default parameters) was used for SSR primer design. Through capillary fluorescence electrophoresis detection, SSR primers with good polymorphism and high amplification success rate were ultimately selected for subsequent data analysis; Perform UPGMA clustering analysis using MEGA and construct a clustering tree. Results A total of 202 361 Unigenes were obtained from the transcriptome database of the needle of T. chinensis var. mairei treated with GA₃, and 12 008 SSR were found by MISA software, which were distributed on 10 894 Unigenes with an occurrence frequency of 5.38% and a mean distance of 7.64 kb per one. Mono-nucleotide repeats were major types, accounting for 51.74% of the total number of SSR, followed by trinucleotide repeats (20.92%) and dinucleotide repeats (20.88%). A/T and AG/CT were dominant repeat motifs, accounting for 49.60% and 10.93% of the total number of SSR repeat types, respectively. A total of 96 SSR primers was selected randomly, and verified with 12 germplasms of T. chinensis var. mairei. 13 pairs of SSR primers with polymorphism were screened out, and the ratio of polymorphic primers was 13.54%. The amplification rate of 13 pairs of primers was as high as 100% in T. cuspidata, but only 23.08% in T. × media. Cluster analysis showed that T. chinensis var. mairei in Taihang Mountain area was clustered into one group, and T. chinensis var. mairei from southern China such as Zhejiang, Fujian and Hunan were clustered into one group, and the genetic distance between the latter three groups was closer. Conclusion The Unigenes information from the transcriptome of T. chinensis var. mairei can be used to identify SSR markers. The SSR markers screened in this study are helpful to the study of genetic diversity, molecular breeding and resource protection of T. chinensis var. mairei.

国家林业和草原局科技发展中心生物安全与遗传资源管理项目（KJZXSA202037）；中央级公益性科研院所基本科研业务费专项资金（CAFYBB2019SY006）