六盘水小黄姜脱毒快繁技术及遗传变异研究  

Study on Virus-free Rapid Propagation and Genetic Variation of Small Yellow Ginger(Zingiber officinale Rosc) in Liupanshui

作　　者：杨松宸 赵德刚[1,3] 赵懿琛 

机构地区：[1]贵州大学农业生物工程研究院,生命科学学院,山地植物资源保护与种质创新省部共建教育部重点实验室,贵阳550025 [2]贵州大学药学院,贵阳550025 [3]贵州省农科院,贵阳550006

出　　处：《分子植物育种》2017年第12期5070-5078,共9页Molecular Plant Breeding

基　　金：贵州省科技计划项目(黔科合LH字[2014]7678)资助

摘　　要：以贵州省六盘水市六枝特区折溪乡小黄姜姜芽为外植体,通过外植体灭菌实验确定了一套高效的姜芽外植体灭菌方法,即剥去姜芽外层组织后,用75%酒精处理30 s,0.1%升汞处理3 min。该方法可在保证存活率为100%时,达到99%的灭菌率。本研究采用正交方法检测不同浓度生长素萘乙酸(1-naphthylacetic acid,NAA)和细胞分裂素6-苄氨基腺嘌呤(6-benzylaminopurine,6-BA)对六盘水小黄姜组培苗生长的影响。数据结果表明,在接种3个月内,正交组平均长出丛生芽1.7个,不定根3.1条。当培养基中激素含量为1.0 mg/L NAA和4.0 mg/L 6-BA时,平均长出丛生芽4.1个,不定根12条。说明该激素浓度组合是促进组培苗丛生芽和不定根生长及分化的最优组合。烟草花叶病毒(tobacco mosaic virus,TMV)酶联检测显示脱毒小黄姜组培苗TMV浓度平均值为0.299 5 ng/m L(未处理小黄姜TMV浓度为3.317 4 ng/m L),说明结合茎尖脱毒与热处理脱毒可以有效除去小黄姜体内的TMV。采用ISSR分子标记技术对小黄姜脱毒苗进行遗传变异分析,从100条ISSR引物中筛选出5条稳定的多态性引物,不同继代小黄姜脱毒苗遗传变异检测表明继代培养8次后的植株与母株的遗传特性均保持一致。A set of efficient ginger bud explant sterilization methods were identified through explant sterilization experiments using Liupanshui small yellow ginger as material, namely stripped bud outer organization and treated with 75% alcohol 30 s, 0.1% Hg Cl23 min. This method could ensure 100% survival rate and reach 99% of the sterilization rate. In this study, the effects of different concentrations of NAA and 6-BA on the growth of ginger tissue culture seedlings were detected using orthogonal method. The data showed that within 3 months after inoculation, the orthogonal group grow cluster buds were at the average of 1.7 and roots 3.1. When the medium hormone content was 0.1 mg/L NAA and 4 mg/L 6-BA, the differentiation and growth of the multiple shoot and adventitious of ginger had a significant advantage, the average of multiple shoot and adventitious were at the average of 4.1 and 12, respectively. Thus, we inferred that this concentration of the hormone could better promotethe differentiation and growth of mu ltiple shoot and adventitious roots on tissue culture seedlings. This study analyzed the influence of different hormone concentration on the differentiation and growth of ginger buds and explored the optimal growth condition of ginger buds explant culture. ELISA detection showed that the average TMV concentration of virus-free seedling was 0.299 5 ng/m L（The untreated seedling TMV concentration was3.317 4 ng/m L）, The results showed that combined with virus free of shoot tip and heat treatment could effectively remove the TMV from the small yellow ginger. ISSR molecular markers were used to analyze the genetic variation of virus-free seedlings of small yellow ginger; five stable polymorphism primers were screened from 100 ISSR primers, genetic variation of different subculture seedlings showed that the genetic characteristic soft and plant were consistent after 8 subcultures.

关 键 词：小黄姜 脱毒培养 正交分析 遗传变异 

分 类 号：S632.5[农业科学—蔬菜学]