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作物学报 ›› 2020, Vol. 46 ›› Issue (12): 1991-1996.doi: 10.3724/SP.J.1006.2020.03025

• 研究简报 • 上一篇    下一篇

玉米雄性不育突变体mi-ms-3的遗传分析及分子鉴定

田士可(), 秦心儿, 张文亮, 董雪, 代明球, 岳兵*()   

  1. 华中农业大学作物遗传改良国家重点实验室, 湖北武汉 430070
  • 收稿日期:2020-05-07 接受日期:2020-08-19 出版日期:2020-08-31 网络出版日期:2020-08-31
  • 通讯作者: 岳兵
  • 基金资助:
    国家重点研发计划项目(2016YFD0100804)

Genetic analysis and characterization of male sterile mutant mi-ms-3 in maize

TIAN Shi-Ke(), QIN Xin-Er, ZHANG Wen-Liang, DONG Xue, DAI Ming-Qiu, YUE Bing*()   

  1. National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, Hubei, China
  • Received:2020-05-07 Accepted:2020-08-19 Published:2020-08-31 Published online:2020-08-31
  • Contact: YUE Bing
  • Supported by:
    National Key Research and Development Program of China(2016YFD0100804)

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摘要：

玉米是杂种优势利用的最好的作物之一, 雄性不育材料作为一种宝贵的种质资源, 对杂种优势的利用具有十分重要的价值。前期筛选得到1个雄性不育突变体, 并将其命名为mi-ms-3。该突变体表现为: 雄穗花药数目减少且不外露, 每个花药只有2个药室, 部分花药退化为膜状并在其末端形成丝状物; 1% I2-KI染色发现花药中含有能正常着色的花粉粒, 与野生型花粉镜检不同之处在于总花粉粒数目的减少; 突变体雌穗花丝增多, 成熟果穗的籽粒两侧各有1个败育的籽粒。通过mi-ms-3与自交系Mo17杂交得到F1完全正常。利用F2分离群体进行遗传分析发现突变表型由隐性单基因控制。利用BSA法, 初步将该基因定位在3号染色体长臂上。随后利用29对SSR标记和10对Indel标记, 将突变基因定位到S-6和umc1027两个标记之间, 物理距离为1.5 cM。该定位区间内有21个候选基因, 通过转座子标签法及测序分析, 最终发现Zm00001d042618 (zmm16)基因ATG上游30 bp处发生了Mu转座子的插入突变。分析表明, mi-ms-3与之前所报道的由于碱基突变造成的sts1突变体的突变方式不同, 是一个新的sts1的等位突变体。RT-PCR分析表明zmm16基因在突变体中表达量降低。mi-ms-3的发现为玉米花器官发育研究以及不育化杂交制种提供了新材料。

关键词: 玉米, 雄性不育, 基因定位, 遗传分析

Abstract:

Maize is one of the best crops in the utilization of heterosis. Male sterile lines are important germplasms for the hybrids production. A male sterile mutant named mi-ms-3 was obtained by screening in a mutator insertion library. The number of male anthers in tassel decreased and not exserted. There were few anthers with only two pollen sacs in the mutant tassels, and some of the anthers were degenerated to membranous and formed filaments at their ends. Although pollens in the anthers could be stained by I2-KI, pollen shedding was abnormal and the number of pollen grains decreased. The number of silks in the ear of the mutant increased, and there was a sterile grain on both sides of the maturated kernel. Fertility of F1 plants, which were obtained by hybridization between mi-ms-3 and maize inbred Mo17, was normal. Genetic analysis of F2 population showed that the mutant phenotype was controlled by a recessive gene. The candidate gene was preliminarily mapped on the long arm of chromosome 3 by BSA and it was located between a SSR marker and an Indel marker with a distance of 1.5 cM. There are 21 candidate genes in this region. It was finally found that the insertion mutation of Mu transposon occurred at 30 bp upstream of the coding region of zm00001d042618 (zmm16) by transponson tagging and sequencing analysis. The results showed that mi-ms-3 was a new allele of sts1, which caused by a single base mutation in the coding region. RT-PCR analysis indicated that the expression of zmm16 in the mutant was decreased. The identification of the new allelic mutant of sts1 in this study would provide new materials for the study of flower development and hybrid seed production.

Key words: maize, male sterile, gene mapping, genetic analysis

表1

基因定位及基因表达分析的引物"

引物名称
Primer name		 正向引物
Forward sequence (5′-3′)		 反向引物
Reverse sequence (5′-3′)		 染色体位置
Chromosome position
umc1973 CAGGCAGAAAAGGAACGGAAC GTGCGAGAGAAGATGGATGATTG 3.05
umc1027 AACTCTGTCTCCGTCACCGTGT GACCTCATCTCGGTGGAAATTG 3.06
S-1 CCGTCATCGACATTTTACTCAG CACAAGAGTTGGTTCGCTG 3.06
S-2 CCTTGTACCCGCTCATCCAT CTCCTACCGACCTGCCTG 3.06
S-3 GTGTTACAAGAACTCAGCGT AGCCTTCGTTCAGTTTCGGT 3.06
S-4 TGACTGGCTTTCGTCCTACT CACTCTCCAGCTGCTTCTCT 3.06
S-5 ACAATTAGCTTAAGAACGC AGCGGTGTTCATGGTACTCA 3.06
S-6 TGGCTACTAGCTAGCTGTGCT CTAAACCTTCACGCACAGCC 3.06
S-7 CCAACTGAGCCGAATCGTTC ATCATCCCATATCCCACGCC 3.06
S-8 GTGGCCCATATGGTCTCAAC CGGCCAGTCGATTAAAGCTC 3.06
S-9 CGACGAAGAACTGACTTGACC TTGCTTGATCATTCCTCCGC 3.06
S-10 GTTTGGGCCGCATTCGAGAGA TATTCTTGCCTCTCCCACCG 3.06
TIR-1 CGGCCTCCATTTCGTCGAATCCCTT
42618 TGCCGGTACTGTGACTGTTC CCTGAGCTCGATCTGCATGG 3.06
42618-2 TCTTTACTCCTCCCCTCCCA CTTCTTGAGGATCCCGTTGC 3.06
q42618 GCCTTAGTGCAGAGATTGACC GCCTTTCCCAGTGCTCCA 3.06

图1

mi-ms-3和野生型表型 A: 野生型(左)和mi-ms-3 (右)雄穗表型, mi-ms-3花药不外露; B1~B2: 花药发育后期, 野生型和mi-ms-3花药横切片, mi-ms-3花药只有2个药室; C1~C4: mi-ms-3突变体小穗, 分别包含0~3个花药; D1~D2: 野生型和mi-ms-3花粉粒; E1: mi-ms-3 (左)和野生型(右)雌穗表型, mi-ms-3雌穗花丝增多; E2: mi-ms-3 (左)和野生型(右)籽粒表型, mi-ms-3每个籽粒对应多个花丝。"

表2

重要重组单株基因型及表型"

umc1973 S-1 S-2 S-3 S-4 S-5 S-6 S-7 S-8 S-9 S-10 umc1027 表型Phenotype
2-2 H H H H A A A A A A A A 突变体Mutant
11-5 H H H H H A A A A A A A 突变体Mutant
9-3 H H H H H H H A A A A A 突变体Mutant
14-8 A A A A A A A A A A A H 突变体Mutant
5-4 H H H H H H H H H H H A 野生型Wild type
20-9 A H H H H H H H H H H H 野生型Wild type

图2

Mu转座子插入突变体的鉴定 1~2: 杂合野生型植株; 3~5: 纯合野生型植株; 6: marker; 7~14: 纯合突变体植株; 15: B73; 16: Mo17。"

图3

zmm16基因序列分析 Mu13转座子插入到zmm16基因ATG上游30 bp处。"

图4

zmm16启动子元件预测分析"

图5

zmm16表达量分析 相对于野生型, mi-ms-3中zmm16基因表达量降低。"

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