无核品种中葡萄18号胚挽救技术体系的建立及其杂交后代早期选择  

Construction of embryo rescue technology system for seedless grape cultivar Zhongputao No.18 and early selection of hybrid offspring

作　　者：陈豆豆 贺亮亮[1] 章鹏[1] 关利平[1] 宋银花[1] 刘三军[1] CHEN Doudou;HE Liangliang;ZHANG Peng;GUAN Liping;SONG Yinhua;LIU Sanjun(Zhengzhou Fruit Research Institute,Chinese Academy of Agricultural Sciences,Zhengzhou 450009,Henan,China)

机构地区：[1]中国农业科学院郑州果树研究所,郑州450009

出　　处：《果树学报》2021年第12期2223-2235,共13页Journal of Fruit Science

基　　金：财政部和农业农村部:国家现代农业产业技术体系;中国农业科学院创新工程(CAAS-ASTIP-2020-ZFRI);河南省科技攻关项目(202102110196)。

摘　　要：【目的】无核品种是葡萄的主要育种目标之一,通过建立无核品种中葡萄18号的胚挽救技术体系,为无核葡萄的胚挽救育种提供参考依据。【方法】通过研究不同胚珠取样时间(盛花后52、53、54、57、58、61和65 d,DAF),培养基类型(NN和ER)及相态(固-液双相和固相),氨基酸(0,2.5 mmol·L^(-1) Ser、Cys、Gln),椰汁(0、100、200 mL·L^(-1)),GA_(3)/KT/NAA组合及4种细胞分裂素TDZ(0和0.2 mg·L^(-1))、6-BA(0和0.2 mg·L^(-1))、KT(0、0.2和0.4 mg·L^(-1))和ZT(0、0.2和0.4 mg·L^(-1))对中葡萄18号胚发育率、胚萌发率或成苗率的影响,并利用无核基因SSR标记对胚挽救杂种苗进行无核性状早期鉴定,初步建立该品种的胚挽救技术体系。【结果】中葡萄18号在DAF 61取样时胚发育率最高(34.33%);基本培养基NN(29.33%)较ER(25%)的胚发育率更高,且固-液双相培养基的效果较固相更佳;添加2.5 mmol·L^(-1) Ser(40.74%)、Cys(43.33%)或Gln(44.21%)的胚发育率均高于对照(29.33%);添加200 mL·L^(-1)椰汁的胚发育率(37.10%)显著高于对照(29.33%)。在3种激素GA_(3)/KT/NAA组合中,发现仅添加GA_(3)和KT时,成苗率最高(60%),而添加NAA的组合均出现异常分化现象,且成苗率较低。四种细胞分裂素TDZ、6-BA、KT和ZT中,添加0.2 mg·L^(-1) ZT更适合胚萌发及成苗,正常成苗的比率达76.36%。分别利用无核SSR标记P3_VvAGL11和5U_VviAGL11对中葡萄18号×玫瑰香的406个杂交株系进行无核性状检测,初步判定229个后代为无核株系,该组合的无核率为56.4%。【结论】中葡萄18号作为胚挽救母本的最佳取样时间为DAF 61前后,可利用固-液双相培养基NN+2.5 mmol·L^(-1) Cys/Gln+5.5 g·L^(-1)琼脂+60 g·L^(-1)蔗糖+2 g·L^(-1)活性炭+0.5 g·L^(-1)水解酪蛋白进行胚珠离体培养,暗培养9~10周后,接种裸胚于WPM+0.2 mg·L^(-1) ZT+20 g·L^(-1)蔗糖+6 g·L^(-1)琼脂+1 g·L^(-1)活性炭进行胚萌发培养。鉴定出拥有无核基因SSR标记的胚挽救苗2【Objective】One of the major objectives of table grape breeding is to produce seedless grapevines.Using stenospermocarpic grapevines as female parents could improve the seedless ratio of offspring via embryo rescue techniques,so they are widely used for breeding seedless grapes.Here,we used the stenospermocarpic grapes Zhongputao No.18 as female parents to breed new seedless grapes and investigated the effects of various factors on the improvement of embryo formation,germination,and plantlet development to construct its embryo rescue technology system,and to provide a reference for the embryo rescue breeding of seedless grapevines.【Methods】The hybrid ovules were obtained from cross combination Zhongputao No.18(seedless grape cultivar)×Muscat Hamburg(seeded muscat-scented cultivar)and the naturally pollinated Zhongputao No.18.In this research,the effects of the stage of ovule development(52,53,54,57,58,61,and 65 days after anthesis),the type of basal medium and its phase,types of amino acids,coconut juice concentrations,and plant growth regulator were assessed on the success rate of the embryo rescue Zhongputao No.18.The gibberellic acid(GA_(3)/),kinetin(KT),and Naphthylacetic acid(NAA)were combined by Uniform Design Experimentation U_(7)(7^(3))to study the effects of the three types of hormones on embryo germination and seedling formation.Moreover,compare the effects of four cytokinins TDZ(0,0.2 mg·L^(-1)),6-BA(0,0.2 mg·L^(-1)),KT(0,0.2 and 0.4 mg·L^(-1))and ZT(0,0.2 and 0.4 mg·L^(-1))on embryo germination and seedling formation.Characters such as development embryo,embryo germination,and seedling formation were recorded.Finally,the molecular marker linked to the seedless gene was employed to detect the seedless hybrids for embryo-rescue seedlings.【Results】When the berries were harvested at 52,53,54,57,58,61,and 65 days after anthesis(DAF),the highest development embryo percentage was observed at 61 days after anthesis,34.33%.Moreover,the ovules were obtained at 16,19,22,26,29,33,36,43,45,51,54,57

关 键 词：无核葡萄 中葡萄18号 胚挽救 技术体系 分子标记辅助选择 

分 类 号：S663.1[农业科学—果树学]