‘赤霞珠’葡萄未成熟合子胚的体细胞胚发生研究  

Somatic embryogenesis in immature zygotic embryos of Vitis vinifera ‘Cabernet Sauvignon’

作　　者：王佳慧 徐伟荣[2,3,4] 郑巧玲 李俊铎 俞沁含 WANG Jiahui;XU Weirong;ZHENG Qiaoling;LI Junduo;YU Qinhan(College of Agronomy,Ningxia University,Yinchuan 750021,China;School of Food&Wine,Ningxia niversity,Yinchuan 750021,China;Engineering Research Center of Grape and Wine,Ministry of Education,Yinchuan 750021,China;Key Laboratory of Modern Molecular Breeding of Dominant and Special Crops in Ningxia,Yinchuan 750021,China)

机构地区：[1]宁夏大学农学院,银川750021 [2]宁夏大学食品与葡萄酒学院,银川750021 [3]葡萄与葡萄酒教育部工程研究中心,银川750021 [4]宁夏优势特色作物现代分子育种重点实验室,银川750021

出　　处：《植物生理学报》2023年第1期67-77,共11页Plant Physiology Journal

基　　金：国家自然科学基金(31860542与32060672);宁夏回族自治区重点研发计划(重大)重点项目(2019BBF02022)。

摘　　要：为建立‘赤霞珠’葡萄(Vitis vinifera ‘Cabernet Sauvignon’)体细胞胚再生体系,以花后80 d未成熟合子胚为外植体,探究了促进初生胚萌发的最佳处理方式以及激素组合对次生体细胞胚直接与间接发生的影响。采用不同浓度(1.0、1.5、2.0、2.5、3.0 g·L^(-1))赤霉素处理、不同时长(2、5、10、15、30 min)超声处理、不同破皮方式(切割种背、切喙+切割种背、磨砂)处理诱导初生胚,进而将已获取初生胚的子叶、胚轴、胚根分别切段放置于添加4种不同浓度的2,4-二氯苯氧乙酸(2,4-D;0.5、1.0、1.5、2.0 mg·L^(-1))和6-苄氨基嘌呤(6-BA;1.0、2.0、3.0、4.0 mg·L^(-1))组合的MS培养基中循环诱导次生胚。结果表明:在初生胚诱导中, 2.5 g·L^(-1)赤霉素、超声2 min、切喙+切割种背三种处理可大幅提高初生胚的诱导率, 11 d时诱导率分别为97.28%、98.64%与92.47%,对照组仅68.42%;在次生胚诱导中,添加0.5、1.0、1.5 mg·L^(-1)2,4-D和1.0、2.0、3.0 mg·L^(-1)6-BA的MS培养基有利于愈伤组织增殖并可通过间接途径产生体胚, 90 d时诱导率在6.45%~17.15%之间。在添加4种不同浓度2,4-D和6-BA组合的MS培养基中, 2.0 mg·L^(-1)2,4-D和4.0 mg·L^(-1)6-BA组合的体细胞胚诱导率最高, 90 d成胚率达到21.33%,次生胚在子叶褐化处直接形成。94%的正常次生胚可萌发并再生植株,生根率达100%。本研究所建立的‘赤霞珠’未成熟合子胚体细胞胚诱导方法为葡萄属植物再生体系优化完善和遗传转化应用研究提供参考依据。To establish a somatic embryo regeneration system for Vitis vinifera ‘Cabernet Sauvignon’, immature zygotic embryos at 80 days after flowering were collected and used as explants. The optimal treatments to promote primary embryo germination and the effects of hormone combinations on the direct and indirect occurrence of secondary somatic embryos were investigated. In order to produce primary embryos,various concentrations of gibberellin(GA3;1.0, 1.5, 2.0, 2.5, and 3.0 g·L^(-1)), durations of ultrasonication(2, 5,10, 15, and 30 min), and techniques of “broken skin”(cutting seed back, cutting beak + cutting seed back,and sandpaper abrasion) were utilized. To develop secondary embryos, MS medium was supplemented with different combinations between 2,4-dichlorophenoxyacetic acid(2,4-D;0.5, 1.0, 1.5, and 2.0 mg·L^(-1)) and 6-benzylaminopurine(6-BA;1.0, 2.0, 3.0, and 4.0 mg·L^(-1)). The secondary embryos were produced by cycling the cotyledons, embryonic axis, and roots of the primary embryos in MS medium supplemented with four different concentration of combinations between 2,4-D(0.5, 1.0, 1.5, and 2.0 mg·L^(-1)) and 6-BA(1.0, 2.0, 3.0, and 4.0 mg·L^(-1)). The result demonstrated that, at the stage of primary embryo induction, 2.5 g·L^(-1)GA3, ultrasonication for 2 minutes, and beak + cutting seed back dramatically improved the induction rate to 97.28%,98.64%, and 92.47%, respectively, compared to 68.42% in the control group. During secondary embryo induction, MS medium with 0.5, 1.0, or 1.5 mg·L^(-1)2,4-D and 1.0, 2.0, or 3.0 mg·L^(-1)6-BA favored callus tissue proliferation and the production of somatic embryos via an indirect pathway, with induction rates ranging from 6.45% to 17.15% at 90 days. The highest induction rate of somatic embryos was achieved in MS medium supplemented with 2.0 mg·L^(-1)2,4-D and 4.0 mg·L^(-1)6-BA, with 21.33% of embryo formation rate at 90 days in MS medium supplemented with four different combinations of concentration, and secondary embryos were formed directly at the

关 键 词：‘赤霞珠’葡萄 未成熟合子胚 体细胞胚 超声处理 直接发生 

分 类 号：S663.1[农业科学—果树学]