彩叶香樟转录组SSR鉴定及指纹图谱构建  

SSR Identification of Transcriptome and Fingerprint Construction in Color-leafed Cinnamomum camphora

作　　者：孔琳琳 孟现东 王建军 俞金健 胡现铬 林二培[1] 黄华宏[1] KONG Lin-Lin;MENG Xian-Dong;WANG Jian-Jun;YU Jin-Jian;HU Xian-Ge;LIN Er-Pei;HUANG Hua-Hong(The State Key Laboratory of Subtropical Silviculture,Zhengjiang Agricultural and Forestry University,Hangzhou 311300,China;Forestry Seed and Seedling Management Station of Zhejiang Province,Hangzhou 311300,China;Ningbo Agricultural Technology Extension Station,Ningbo 315012,China)

机构地区：[1]浙江农林大学亚热带森林培育国家重点实验室,杭州311300 [2]浙江省林业种苗管理总站,杭州311300 [3]宁波市农业技术推广总站,宁波315012

出　　处：《农业生物技术学报》2023年第11期2423-2434,共12页Journal of Agricultural Biotechnology

基　　金：浙江省省院合作林业科技项目(2021SY02);杭州市科技发展计划项目(2020ZDSJ0909);浙江省农业(林木)新品种选育重大科技专项课题(2021C02070-7-4)。

摘　　要：彩叶香樟(Cinnamomum camphora)是近年选育的春色叶新品种,市场前景广阔,但尚未建立有效的鉴别技术体系,影响了新品种权保护及市场流通。为了实现彩叶香樟新品种的快速鉴定,本研究基于转录组开发SSR标记,并构建其指纹图谱。利用Illumina高通量测序平台对彩叶香樟新品种’涌金’(C.camphora’Yongjin’)进行转录组测序,共获得129175条单基因簇(Unigene),总长度为92429171 bp。通过MISA(MIcroSAtellite identification tool)软件对转录组序列进行SSR搜索,发现28895条序列中含有36430个SSR位点,平均2537.2 bp出现一个SSR,发生频率为22.37%;单核苷酸、二核苷酸、三核苷酸重复单元分别占总SSR的49.74%、29.12%和19.93%。随机选择54对引物进行PCR扩增,能够清晰扩增出期望产物的引物有39对,其中32对在23份香樟种质资源中表现多态性。采用非加权组平均法(unweighted pair-group method with arithmetic means,UPGMA)对23份香樟种质进行聚类分析,其相似系数为0.69~0.88。在相似性系数0.70处,可将供试材料分为三大类群。选出c117334、c90845、c108367、c115142、c49114、c101928共6对引物组合构建指纹图谱,能有效将3个新品种与其子代以及不同地理来源的香樟种质完全区分。本研究为彩叶香樟新品种鉴定、知识产权保护以及分子辅助育种提供技术支持和理论依据。Color-leafed Cinnamomum camphora is an ornamental plant variety with colored leaves in spring.Although these varieties have broad market prospects,a technology system for distinguishing them effectively has not yet been established,which affects the protection of new variety rights and market circulation.In this study,in order to rapidly identify 3 color-leafed varieties,their fingerprints were constructed using SSR markers derived from the transcriptome of C.camphora.The transcriptome of C.camphora'Yongjin'was sequenced using the Illumina high-throughput sequencing platform,and a total of 129175 Unigenes were obtained,with a total length of 92429171 bp.The SSR search of the transcriptome sequences by MIcroSAtellite identification tool(MISA)revealed that 28895 sequences contained 36430 SSR sites,with an average of 1 SSR per 2537.2 bp and a frequency of 22.37%.Mononucleotide repeats(accounting for 49.74%)were the most abundant,followed by dinucleotide(29.12%)and trinucleotide(19.93%).Fifty-four primer pairs were randomly selected for PCR amplification,and 39 of them were successfully amplified.In total,32 primer pairs were polymorphic in 23 C.camphora germplasms.In addition,unweighted pair-group method with arithmetic means(UPGMA)cluster analysis showed that the similarity coefficients of 23 germplasms ranged from 0.69 to 0.88.At the similarity coefficient of 0.70,these germplasms could be classified into 3 major groups.Finally,6 primer pairs(c117334,c90845,c108367,c115142,c49114 and c101928)were selected to construct the fingerprints,which could effectively distinguish the 3 new varieties from their offsprings and other germplasms from different geographical sources.The results could provide technical support and theoretical basis for new variety identification,intellectual property protection and molecular assisted breeding of C.camphora.

关 键 词：香樟 彩叶新品种 转录组 SSR 指纹图谱 

分 类 号：S792.23[农业科学—林木遗传育种]