花椒转录组SSR信息分析及其分子标记开发  

Analysis of SSR Loci and Development of Molecular Markers in Zanthoxylum bungeanum Transcriptome

作　　者：候丽秀 魏安智[1] 王丽华[2] 刘玉林[1] HOU Li-Xiu;WEI An-Zhi;WANG Li-Hua;LIU Yu-Lin(College of Forestry, Northwest A＆F University, Yangling 712100, China;Institute of Biotechnology and Seed, Sichuan Academy of Forestry Science, Chengdu 610081, China)

机构地区：[1]西北农林科技大学林学院,杨凌712100 [2]四川省林业科学研究院生物技术与良种研究所,成都610081

出　　处：《农业生物技术学报》2018年第7期1226-1236,共11页Journal of Agricultural Biotechnology

基　　金：西北农林科技大学博士科研启动基金(No.2452015296);国家林业局林业公益性行业科研专项(No.201304706)

摘　　要：匮乏的基因组信息是导致花椒(Zanthoxylum bungeanum)分子标记缺乏的根本原因。本研究通过高通量测序技术结合MISA软件分析花椒果皮转录组中SSR的分布类型及特点,以来源于中国6个省份的33份花椒种质对随机合成的80对引物进行多态性筛选,继而开发表达序列标签SSR(expressed sequence tags-SSR,EST-SSR)标记并分析不同花椒种质的遗传多样性。结果显示,在转录组测序获得的163 340条Unigenes中,共筛选到27 905个SSR位点,分布于23 422条Unigenes中,SSR发生频率和平均分布距离分别为14.34%和2.94 kb。其中,1 110个SSR位于编码区序列(coding sequence,CDS);4 358和3 856个SSR分别位于5'端非编码区(5-untranslated regions,5'-UTR)和3'端非编码区(3'-untranslated regions,3'-UTR)。单、二和三核苷酸为优势重复类型,分别占SSR总数的66.48%、17.37%和14.89%,且A/T、AG/TC和AAG/TTC分别是单、二和三核苷酸的优势重复基元。此外,本研究成功设计12 478对SSR特异位点引物。在随机选取的80对引物中,共有15对表现出多态性。多态性信息含量(polymorphism information content,PIC)、观测等位基因数(observed number of alleles,Na)、有效等位基因(effective number of alleles,Ne)、观测杂合度(observed heterozygosity,Ho)、期望杂合度(expected heterozygosity,He)、Shannon's指数(Shannon's information index,I)和Nei's基因多样性指数(Nei's gene diversity index,H)的平均值分别为0.50、3.27、2.43、0.69、0.58、0.97和0.57。利用UPGMA聚类,可将33份供试花椒种质分为4类,与供试种质地理位置划分基本保持一致。本研究为花椒种质资源鉴定、分子标记辅助选择育种及遗传多样性分析等提供标记资源。The short of genomic information is the fundamental cause which makes the lack of molecular markers in Zanthoxylum bungeanum, an aromatic plant of the family Rutaceae which native to China. In this study, MISA software was used to screen and analyze the distribution types and characteristics of simple sequence repeats （SSRs） in the pericarp transcriptome of Z. bungeanum which obtained by high-throughput sequencing technology. Then, the SSR primer pairs were designed by Primer 3.0 software and 80 primer pairs of them were selected randomly to test polymorphism among 33 individuals which from six provinces of China to develop the EST-SSR markers and analyzed the genetic diversity of Z. bungeanum. As a result, 163 340 unigenes which mainly distributed in the length interval from 200 bp to 400 bp were obtained from transcriptome sequencing and 27 905 SSRs included mono-, di-, tri-, tetra-, penta-, and hexanucleotide motifs, which located in 23 422 unigenes were identified. The frequency of occurrence and mean distribution distance of SSRs were 14.34% and 2.94 kb, respectively. The most abundant type was mono-nucleotide repeat motifs, followed by di-, tri-, tetra-, penta-, and hexanucleotide repeat motifs, which accounting for 66.48% （18552）, 17.37% （4848）, 14.89% （4155）, 1.08% （301）, 0.07% （20） and 0.10% （29） of the total SSR number, respectively. A/T, AG/TC, AAG/TTC and AAAT/TTTA which represented 98.94%, 47.11%, 23.01% and 28.24% of the total number for mono-, di-, tri- and tetranucleotide motifs were the dominant motif identified in each repeat motifs, respectively. According to the result of coding sequence （CDS） prediction, 1 110, 4 385 and 3 856 SSR loci were detected which located in CDS, 5＇-untranslated regions （5＇-UTR） and Y-untranslated regions （3＇-UTR）, respectively. However, the remaining 18 581 SSRs were undetermined due to there was not enough information to delimit the SSR loci in the CDS, 5＇-UTR and Y-UTR regions of these unigenes. For the determined loc

关 键 词：花椒 转录组 简单重复序列(SSR)标记 遗传多样性 

分 类 号：S565.9[农业科学—作物学]