棉花黄萎病菌T-DNA插入突变体库的构建及变异性状的分析  

Construction of T-DNA Inserted Transformation Library of Verticillium dahliae on Cotton and Mutative Traits of the Mutants

作　　者：周芳芳[1] 李志芳[1] 冯自力[1] 赵丽红[1] 师勇强[1] 朱荷琴[1] 

机构地区：[1]国家棉花生物学重点实验室,中国农业科学院棉花研究所,河南安阳455000

出　　处：《西北农业学报》2012年第8期19-25,共7页Acta Agriculturae Boreali-occidentalis Sinica

基　　金：转基因生物新品种培育重大专项(2011ZX08011-002);中央级公益性科研院所基本科研业务费专项2012年度所长基金项目(SJB1203);科技基础性工作专项(SB2007FY027)

摘　　要：利用农杆菌介导遗传转化(ATMT)的方法,成功构建含2 000个转化子的棉花黄萎病菌强毒菌株Vd080的突变体库。200μmol/L乙酰丁香酮(AS)作诱导剂,25℃共培养48h,棉花黄萎病菌孢子量为106 mL-1,其转化效率达150～540个转化子。进一步研究发现,供试突变体的T-DNA成功插入Vd080基因组,且均为单拷贝插入,转化子的潮霉素B基因能够稳定遗传。309个突变体中,53.7%的突变体菌落形态与初始菌株Vd080相同,均产生黑色的微菌核,不产生黑色素微菌核的菌丝型仅占17.5%。与初始菌株相比,随机选取的85个突变体,产孢量、生长速率、粗毒素分泌量及致病力发生显著变异的菌株分别占36.4%、12.9%、50.6%和29.4%,四者之间不存在明显的相关性。A mutant library, which intiated from a high virulent Verticillium dahliae strain Vd080 and contained 2 000 mutants, was constructed by Agrobacterium-mediated transformation. 200 μmol/L Acetosyringone and 48 hours of co-cultivation period at 25 ℃ were optimal for the trans- formation and reached 150-540 transformants per 106 spores. The results of further study indicated that T-DNA of tested mutants inserted into the genome of Vd080 successfully, and T-DNAs in 6 mutants selected randomly were all single copy. Besides, all of the tested transformants maintained their resistance to hygromycin B. Like wild strain Vd080, 53.7%mutants produced black microsclerotia in their colonies. 17.5% mutants produced white or yellow mycelia without black microsclerotia. Mutants which exhibited significantly different from the wild strain Vd080 on spore yield, growth rate,crude toxin and pathogenicity were 36.4%, 12.9%, 50.6% and 29.4%, respectively. There was no correlation among the four parameters.

关 键 词：农杆菌介导转化 棉花黄萎病菌 突变体库 变异性状 

分 类 号：S435.621[农业科学—农业昆虫与害虫防治]