绿豆EMS诱变突变体库的构建及表型分析

doi:10.11924/j.issn.1000-6850.casb19020066

摘要/Abstract

摘要：

为探索适于绿豆突变体库构建的最佳甲基磺酸乙酯(EMS)浓度和处理时间,以‘皖科绿3号’种子为研究材料,设置4种EMS浓度（0.6%,1.0%,1.4%和1.8%）和4种时间（6 h,10 h,14 h和18h）共16个组合,并以半致死剂量为突变体库构建的条件。结果表明,‘皖科绿3号’的最佳诱变条件为1.0%/10 h。用该条件处理6000粒成熟的‘皖科绿3号’种子,获得M₁代植株998份。对M₂代群体中植株叶色、叶形、株高、花器官、开花期等性状进行鉴定。共发现表型变异材料324株,变异频率为2.97%。突变体中叶色变异和矮秆是主要的变异类型,分别占植株总数的1.10%和0.46%。利用基因组重测序技术对4份突变材料的突变位点进行鉴定,测序结果表明碱基突变频率为1/37.78 kb。实验确定‘皖科绿3号’的最佳诱变条件为1.0%/10 h。研究结果也表明利用EMS诱变的方法可获得丰富的表型变异材料,可用于绿豆品种遗传改良和功能基因组学研究。

关键词: 绿豆, 甲基磺酸乙酯, 突变体库, 表型, 测序

Abstract:

To find out the optimal concentration and treatment time of ethyl methyl sulfonate (EMS) for mung bean mutant library construction, ‘Wankelü 3’ was used as material, 16 treatment combinations including 4 EMS concentrations (0.6%, 1.0%, 1.4% and 1.8%) and 4 kinds of treatment time (6, 10, 14 and 18 h) were employed to treat the seeds, and the semi-lethal dose was selected for mutant library construction. The results showed that the optimal condition for ‘Wankelü 3’ mutagenesis was 1.0%/10 h. 6000 mature seeds of ‘Wankelü 3’ were treated under this optimal condition, 998 M₁ lines were obtained. Traits such as leaf color, leaf shape, plant height, flower morphology and flowering time of M₂ generation plants were investigated. A total of 324 mutants were obtained, and the mutation frequency was 2.97%. Among these mutants, leaf color mutation and dwarf were the dominated variation types, accounting for 1.10 % and 0.46%, respectively. Mutation sites of 4 mutant materials were determined by genome re-sequencing, the result showed that their base mutation frequency was 1/37.78 kb. These results also demonstrated that using EMS mutagenesis method, abundant phenotypic variant materials could be obtained, which could be used in genetic improvement and genome functional analysis for mung bean.

Key words: mung bean, ethyl methane sulfonate (EMS), mutant library, phenotype, sequencing

中图分类号:

S322

叶卫军, 杨勇, 张丽亚, 田东丰, 张玲玲, 周斌. 绿豆EMS诱变突变体库的构建及表型分析[J]. 中国农学通报, 2020, 36(17): 36-41.

Ye Weijun, Yang Yong, Zhang Liya, Tian Dongfeng, Zhang Lingling, Zhou Bin. Construction of EMS Mutagenesis Mung Bean Mutant Library and Phenotypic Analysis[J]. Chinese Agricultural Science Bulletin, 2020, 36(17): 36-41.

图/表 4

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性状	变异类型	突变体株数	突变频率/%
叶片	叶色	120	1.10
叶片	叶型	32	0.29
茎	高秆	12	0.11
	矮秆	50	0.46
	畸形茎	21	0.19
	茎色	3	0.03
花	畸形花	4	0.04
花期	早花	19	0.17
花期	晚花	25	0.23
育性	半不育	32	0.29
育性	全不育	15	0.14
荚	长荚	6	0.06
荚	短荚	25	0.23
合计		364	3.34
复合性状突变		40	0.37
实际数量		324	2.97

性状	变异类型	突变体株数	突变频率/%
叶片	叶色	120	1.10
叶片	叶型	32	0.29
茎	高秆	12	0.11
	矮秆	50	0.46
	畸形茎	21	0.19
	茎色	3	0.03
花	畸形花	4	0.04
花期	早花	19	0.17
花期	晚花	25	0.23
育性	半不育	32	0.29
育性	全不育	15	0.14
荚	长荚	6	0.06
荚	短荚	25	0.23
合计		364	3.34
复合性状突变		40	0.37
实际数量		324	2.97

编号	测序深度/×	测序总碱基数/bp	突变碱基数/bp	突变频率/1/kb
ME1	18.33	7883350874	161372	48.85
ME2	20.15	8666094933	156116	55.51
ME3	18.20	7827440585	157308	49.76
ME4	17.85	7664010506	161813	47.36
平均	18.63	8010224225	159152	50.37

编号	测序深度/×	测序总碱基数/bp	突变碱基数/bp	突变频率/1/kb
ME1	18.33	7883350874	161372	48.85
ME2	20.15	8666094933	156116	55.51
ME3	18.20	7827440585	157308	49.76
ME4	17.85	7664010506	161813	47.36
平均	18.63	8010224225	159152	50.37