利用InDel标记鉴定不同橘柚品种  

Identification of tangelo varieties with InDel markers

作　　者：余歆 郭意 刘小丰[1] 万润楚 江东[1] 曹立[1] YU Xin;GUO Yi;LIU Xiaofeng;WAN Runchu;JIANG Dong;CAO Li(Citrus Research Institute,Southwest University,Chongqing 400712,China;Sichuan Dongpo Chinese Paocai Industrial Technology Research Institute,Meishan 620036,Sichuan,China)

机构地区：[1]西南大学柑桔研究所,重庆400712 [2]四川东坡中国泡菜产业技术研究院,四川眉山620036

出　　处：《果树学报》2024年第8期1477-1489,共13页Journal of Fruit Science

基　　金：柑橘种质资源精准鉴定项目(19230691);抚州市“揭榜挂帅”项目(XMBH00091)。

摘　　要：【目的】橘柚(tangelo)是一类含有橘(mandarin,Citrus reticulata Blanco)和柚[pummelo,C.maxima(Burm.)Merr.]血统的杂种柑橘。旨在挖掘能快速有效区分和鉴定不同橘柚品种的插入缺失(InDel)分子标记。【方法】利用第二代测序技术对两种橘柚(阳光1号和阳光2号)及其亲本(爱媛28号和春香)进行全基因组重测序,通过生物信息学分析和琼脂糖凝胶电泳检测,挖掘、筛选和确定大于30 bp或小于-30 bp的InDel标记,使用这些标记鉴定和区分10个不同橘柚品种,并对共51份种质资源进行基因分型及遗传多样性分析。【结果】在4个柑橘品种的基因组共鉴定到607376个InDel位点,产生657414种变异方式,并进一步筛选出48个大于30 bp或小于-30 bp的InDel标记。他们适用于普通琼脂糖凝胶电泳检测,操作简单,省力省时,成本低。这些InDel标记能够对10个橘柚品种进行基因分型,区别和鉴定不同橘柚品种,在51份种质资源中具有遗传多样性,其中I3_744、I7_170、I8_516、I8_533、I9_669等InDel标记具有高多态信息含量(PIC)和期望杂合度(He)及低最大等位基因频率(MAF)的特点。基于这48个InDel标记基因分型的主坐标分析(PCoA)展示了51份种质资源的遗传距离,他们在PCoA二维散点图中的分布与已知的柑橘类型划分总体一致。【结论】发掘的48个适合普通琼脂糖电泳检测的InDel标记能准确快速区分和鉴定不同橘柚品种,也有助于对柑橘作物的遗传多样性和群体进行分类分析。【Objective】Tangelo is a type of citrus hybrid that has mandarin(Citrus reticulata Blanco)and pummelo[C.maxima(Burm.)Merr.]pedigrees.Owing to the characteristic blended flavors of mandarin and pummelo,tangelo is increasingly favoured by consumers,and receiving attentions from citrus breeders.Besides the conventional tangeloes such as Sweet Spring and Haruka,previously we have developed two elite tangelo varieties called Yangguang No.1 and Yangguang No.2 derived from a crossing of Ehime Kashi No.28 and Haruka.At present,the cultivated area of Yangguang No.1 is expanding.Precise and efficient variety identification is crucial for protecting plant variety rights.Molecular markers,such as SNP,SSR and InDel,have been broadly applied for genotyping,variety identification,genetic diversity analysis and pre-selection in various crops.This study aimed to discover whole-genome InDels by comparison of whole-genome sequencing data of Yangguang No.1,Yangguang No.2 and their parents,and developed agarose gel-resolved InDel markers for genotyping and discriminating different tangelo varieties.Furthermore,the application of the selected InDel markers for genetic analysis of a wider range of citrus germplasm resources was also investigated.【Methods】The plant materials for whole genome re-sequencing were the healthy leaves of Yangguang No.1 and Yangguang No.2,and their female parent Ehime Kashi No.28 and male parent Haruka.Ten tangelo varieties were tested for the genotyping,discrimination and identification with the selected InDel markers.Altogether,20 mandarins,10 tangeloes,2 pummeloes,5 sweet oranges[C.sinensis(L.)Osbeck],2 citrons(C.medica L.),3 lemons[C.limon(L.)Burm.f.],2 yuzues(C.junos Siebold.ex Tanaka),2 grapefruits(C.paradisi Macf.),2 kumquats(Fortunella spp.),3 trifoliate oranges(Poncirus trifoliata Raf.)were used for genetic diversity analysis.The whole-genome re-sequencing was carried out with NovaSeq 6000 Sequencer(Illumina Inc.,San Diego,USA)in Biomarker Technologies(Beijing,China).The clean reads were mapped

关 键 词：柑橘 橘柚 INDEL 分子标记 遗传多样性 

分 类 号：S666[农业科学—果树学]