摘要： 本研究从00余株灰葡萄孢ATMT转化子中筛选到产孢缺陷菌株BCG-183.利用CTAB法提取野生型和BCG-183的基因组DNA,Southern杂交证实T-DNA标记在突变菌株中为单拷贝插入;对该菌株表型进行分析,发现BCG-183菌株菌落呈白色,菌丝产量较野生型大;致病性较野生型强;利用血球计数板计算产孢量,表明BCG-183不产生分生孢子;胞壁降解酶活性比野生型强.利用TAIL-PCR技术确定T-DNA插入位点侧翼序列,生物信息学分析表明,T-DNA插入到灰葡萄孢基因组超级重叠群2(Supercontig 2)中假定蛋白编码基因(BClG_0755)的3'末端.RT-PCR结果表明,BCJG_0755基因在突变菌株中没有表达.推测BC1G_0755可能与灰葡萄孢产孢相关.

关键词: 灰葡萄孢, 产孢量, TAIL-PCR, RT-PCR

Abstract: Conidiation-deficiency transformant BCG-183 was obstained from more than 400 ATMT transformants.The genomic DNA of wild type and BCG-183 were extracted by CTAB method,and southern blot showed that one copy of T-DNA had inserted into the mutant's DNA.By analysing the phenotype of the mutant,we found that mutant's hyphae were more dense than the wild type,while its colony was white.The mutant didn't produce conidium.And the cell-wall-degrading enzyme activities were stronger than that of the wild type′s.T-DNA insertion site flanking sequence was amplified by TAIL-PCR.It showed that the T-DNA had inserted into the putative protein coding genes(BC1G_07455)3′termination,which localized in the Supercontig 42(Supercontig42)of Botrytis cinerea genome.RT-PCR results showed that BC1G_07455 didn′t express in the mutant.These results indicated that BC1G_07455 may involve in the conidiation of Botrytis cinerea.

Key words: Botrytis cinerea, Conidiation, TAIL-PCR, RT-PCR

中图分类号: 

• S36.631