基于SSR荧光标记的杧果种质资源遗传多样性分析及分子身份证构建

Genetic Diversity Analysis and Molecular ID Construction of Mango Germplasm Based on SSR Fluorescence Markers

丰富的种质资源是杧果新品种选育和产业发展的基础。为保护和利用杧果种质资源,本研究利用课题组前期开发的TP-M13-SSR标记对杧果种质资源保护广西创新基地圃内保存的145份杧果地方品种、育成品种及其近缘野生种进行遗传多样性分析和分子身份证构建。结果表明:12对引物的平均观测等位基因数为3.2838、平均观察杂合度(H_(o))为0.5858、平均期望杂合度(H_(e))为0.6725、平均Shannon指数(I)为1.3383、平均Nei基因多样性指数(N_(a))为0.6702、多态信息含量(PIC)分布范围在0.5036~0.7827之间,平均值为0.6396,所有引物为高度多态性位点,说明TP-M13-SSR荧光引物可以为杧果的遗传多样性分析提供研究数据;145份材料的遗传相似性系数变化范围为0.5676~1.000,平均为0.7417,其中爱文与印度杧1号遗传相似性系数为1.000,扁桃杧田阳20-2与大头香杧、扁桃杧田阳20-2与硕帅杧、金煌杧与桂热杧10-1的遗传相似性系数最小,均为0.5676;在遗传相似性系数为0.7060时,145份种质分为2个类群,I类群包括108份杧果和20份扁桃,种质数量最多,占总数的88.9%,Ⅱ类群包括17份材料,全部为扁桃。在遗传相似性系数为0.7330时,I类群可进一步分为5个亚群,其中I-1和I-3亚群种质数量最多,占所有杧果的91.92%,UPGMA聚类分析表明扁桃未严格按照种属关系聚在一起,杧果的整体聚类结果与其地理来源基本一致;对145份材料的扩增产物进行SSR荧光标记毛细管电泳检测获得指纹图谱,采用数字和字母相结合的编码方式获得分子身份证,通过分子身份证进行种质鉴定,每一对引物可平均区分12.4份种质,鉴定率明显高于前人研究,表明TP-M13-SSR检测技术比目前广泛采用的变性聚丙烯酰胺凝胶电泳检测技术在杧果种质鉴定上更具优势。本研究结果为杧果及其近缘种种质资源的收集整理和新品种的选育提供科学依据。

Abundant germplasm resources are the basis for mango variety breeding and industrial development.For better protection and utilization of mango germplasm resources,the TP-M13-SSR marker developed by our team previously were used to analyze the genetic diversity and construct molecular ID of 145 mango germplasms containing local cultivars,bred varieties and wild relative species,which stored in the nursery of Guangxi Innovation Base of mango germplasm resources conservation.The results showed that the average number of observed alleles for the 12primer pairs was 3.2838,the average observed heterozygosity (H_o) was 0.5858,the average expected heterozygosity (H_e)was 0.6725,the average Shannon index (I) was 1.3383,and the average Nei’s gene diversity index (N_a) was 0.6702.The polymorphism information content (PIC) of the 12 primer pairs ranged from 0.5036 to 0.7827,with an average value of0.6396.All the primers were highly polymorphic sites.The result suggested that TP-M13-SSR primer could provide data support for genetic diversity analysis of mango.The genetic similarity coefficient of 145 materrials varied from0.5676 to 1.000,with an average of 0.7417.The genetic similarity coefficient between Irwin and Indian No.1 was 1.000.The minimum genetic similarity coefficient was 0.5676,between M.persiciformis 20-2 and Dadouxiang mango,M.persiciformis 20-2 and Shuoshuai mango,Jinhuang mango and Guire 10-1 mango.All the 145 mango germplasms were divided into two groups when the genetic similarity coefficient was 0.7060.Group I contained 108 mango species and20 M.persiciformis species,accounting for 88.90%of the total number of germplasms.GroupⅡcontained 17 specimens,all of which were M.persiciformis species.Group I could be further divided into five subgroups when the genetic similarity coefficient was 0.7330,among which subgroups I-1 and I-3 were the most abundant,accounting for 91.92%of all mango germplasms.The results of UPGMA clustering analysis showed that M.persiciformis were not clustered strictly according to the species relationship,and the overall clustering result of mango germplasms was basically consistent with its geographical origin.All the 145 materials were amplified by 12 pairs of SSR fluorescent primers to obtain the fingerprint map,and the molecular ID was obtained by the assignment of numbers and letters combination.Each pair of primers could distinguish 12.4 germplasms on average,and the identification rate was significantly higher than that of previous studies,indicating that TP-M13-SSR had more advantageous than denaturing polyacrylamide gel electrophoresis in mango germplasm identification.This study would provide scientific basis for the collection and utilization of germplasm resources and variety breeding of mango.It is also proposed for molecular identification of bred varieties,which is of great significance to the development of mango industry for providing methods for molecular identification and intellectual property of bred varieties.