谷瘟病菌无毒基因PWL家族检测及变异分析

doi:10.7668/hbnxb.20190568

摘要/Abstract

摘要： 为了确定不同地区谷瘟病菌群体中无毒基因PWL家族的分布和变异情况，从全国谷子主产区的不同区域采集分离了252株谷瘟病菌单孢菌株，提取基因组DNA，参考目前已知稻瘟病菌中无毒基因PWL家族的核苷酸序列开发合成了5对引物，通过PCR扩增对252株谷瘟病菌进行PWL家族基因检测，并对扩增片段进行测序分析。结果表明，252个谷瘟病菌菌株都不包含PWL1基因，65个菌株包含出PWL2基因，扩增率25.8%，252个菌株都包含PWL3与PWL4基因，扩增率100%。谷瘟病菌PWL3基因与稻瘟病菌中同源基因相比，在基因编码区存在849 bp的碱基插入，插入片段与non-LTR retrotransposon：MGL的相似度为96.7%，利用此差异可通过PCR技术快速区分谷瘟病菌与稻瘟病菌。谷瘟病菌PWL2基因存在多处单核苷酸变异，通过分析将其划分14个单倍型，分别为P1~P14。单倍型P1占绝对优势包含46个菌株，以此为基础衍生出其他主要单倍型，其次为单倍型P12包含7个菌株，并且多地区都存在特异性单倍型，说明我国谷瘟病菌种群具有较高的遗传多样性。谷瘟病菌中不存在PWL1无毒基因，PWL2无毒基因仅存在部分菌株中，PWL3和PWL4无毒基因存在所有谷瘟菌株中，PWL3和PWL4无毒基因对应的抗病基因在谷子上有重要的应用价值。

关键词: 谷子, 谷瘟病菌, PWL基因家族, 基因克隆, 无毒基因

Abstract: To identify the distribution and variation of the avirulence gene PWL family in 252 single spore Magnaporthe oryzae which was isolated from foxtail millet in different regions, a total of 252 strains of M. oryzae were isolated from different regions of millet growing area in China,and then the genomic DNA of 252 M. oryzae strains was extracted for PCR amplification.Five pairs of primers for PCR were designed and synthesized according to the homologous PWL genes sequence in Magnaporthe grisea. The PCR products were sequenced to analyze the distribution of PWL gene family in M. oryzae strain. The results showed that PWL1 gene was not detected in all isolates, and 65 isolates detected PWL2 gene (amplification rate was 25.8%). Avirulence genes PWL3 and PWL4 were found in all isolates of M. oryzae (amplification rate was 100%). Meanwhile,compared with the homologous genes in M. grisea, there was an 849 bp insertion in the gene coding region of PWL3 gene in all isolates from foxtail millet, which could be used to distinguish these two fungi quickly by PCR detection. And the similarity between the insertion and non-LTR retrotransposon:MGL was 96.7%.According to the abundant single nucleotide variants in the PWL2 gene, the 252 M. oryzae strains were divided into 14 haplotypes, named P1-P14 respectively. Among the 14 haplotypes,haplotype P1 was dominant haplotype which contained 46 strains, and the second one was P12 which contained 7 strains. Moreover, specific haplotype had been found in many regions, these results suggested that M. oryzae had high genetic diversity in China. PWL1 gene was not found in M. grisea from foxtail millet and only partial isolates contained PWL2 gene. The avirulence genes PWL3 and PWL4 widely distributed in all isolates,so the blast resistance gene corresponding to the PWL3 and PWL4 avirulence gene could be used to control the foxtail millet blast by disease resistant breeding.

Key words: Foxtail millet, Magnaporthe oryzae, PWL gene family, Gene clone, Avirulence gene

中图分类号:

S432.4+1

申浩冉, 白辉, 任世龙, 王璐, 王永芳, 全建章, 董志平, 李志勇. 谷瘟病菌无毒基因PWL家族检测及变异分析[J]. 华北农学报, 2020, 35(3): 178-183. doi: 10.7668/hbnxb.20190568.

SHEN Haoran, BAI Hui, REN Shilong, WANG Lu, WANG Yongfang, QUAN Jianzhang, DONG Zhiping, LI Zhiyong. Detection and Variation Analysis of PWL Gene Family of Magnaporthe oryzae in Foxtail Millet[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2020, 35(3): 178-183. doi: 10.7668/hbnxb.20190568.

http://www.hbnxb.net/CN/Y2020/V35/I3/178

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