Identification and Genetic Analysis of Waxy Sparse Mutant wax1 in Rice

doi:10.16819/j.1001-7216.2019.9101

Abstract

Abstract:

【Objective】 Wax is an important protective substance on plant surface. Screening and identification of waxy-related mutants help elucidate the genetic mechanism of waxy synthesis in rice. 【Method】A waxy sparse mutant, waxy1, was isolated from an ethylmethylsulfone (EMS) mutagenic population of Xiangzaoxian 6. The morphological characteristics and agronomic traits of waxy1 mutant were analyzed. The F₂ segregating population was constructed from a cross between waxy1 and japonica variety 02428, and the recessive individuals were selected for fine mapping of WAXY1 gene. In addition, the expressions of related genes were analyzed by quantitative PCR. 【Results】Compared with the wild type, wax1 mutant showed many morphological abnormalities, including small wrinkled leaves, shorter panicle and reduced waxy crystals on leaf surface. The wax1 mutant was also featured by increased effective panicle number, and obviously decreased plant height, 1000-grain weight and number of grains per panicle. Genetic analysis implied that the mutant phenotype was controlled by a pair of recessive nuclear gene. The WAXY1 gene was fine-mapped to a 49.8 kb interval between markers RM5806 and P1 on the chromosome 10. A total of 4 ORFs (open reading frames) were found in this region. Sequencing analysis indicated that a single base transition was present in the coding region of 3-ketoacyl-CoA synthase, which resulted in an amino acid substitution. In addition, mutation of WAXY1 increased the expression of KNOX gene OSH6, and caused differential expressions of some IAA genes. 【Conclusion】The WAX1 gene mutation resulted in the decrease of waxy crystals on leaf surface and abnormalities in plant growth and development through affecting shoot apex meristem and auxin signal transduction.

Key words: rice, waxy, gene mapping, auxin

摘要：

【目的】蜡质是植物表面的一种重要保护物质,筛选和鉴定水稻蜡质相关突变体有助于解析水稻蜡质形成的遗传机制。【方法】利用EMS诱变籼稻品种湘早籼6号,从突变体库中筛选出一个蜡质稀少的突变体wax1,考查突变体的形态特征和农艺性状,利用突变体与02428杂交的F₂群体定位目标基因,并通过荧光定量PCR分析相关基因的表达情况。【结果】与野生型相比,wax1突变体具有叶片短小皱褶、叶表面蜡质晶体减少、穗长变短等形态特征;在农艺性状方面,突变体的株高、每穗总粒数和千粒重显著降低,但有效穗数显著高于野生型;遗传分析表明,wax1的突变表型受一对隐性核基因控制,将WAX1基因定位在第10染色体上SSR标记RM5806与InDel标记P1之间,物理距离约为49.8 kb;定位区间内的测序结果表明,突变体中β-酮脂酰-CoA合酶的编码基因发生单碱基突变,导致催化活性中心的一个氨基酸发生改变。WAX1基因的突变显著提高了同源异型盒基因OSH6的表达,同时引起部分IAA基因的差异表达。【结论】WAX1基因突变引起叶表面蜡质晶体的减少,同时通过影响茎尖分生组织和生长素的信号转导引起植株生长发育等多方面的异常。

关键词: 水稻, 蜡质, 基因定位, 生长素

CLC Number:

Q755
S511.01

Xiaowu PAN, Yongchao LI, Wenqiang LIU, Haibo XIONG, Zheng DONG, Dan XUE, Xinnian SHENG, Wenjin ZHAO, Xiucai WEI, Xiaoxiang LI. Identification and Genetic Analysis of Waxy Sparse Mutant wax1 in Rice[J]. Chinese Journal OF Rice Science, 2020, 34(1): 1-7.

潘孝武, 黎用朝, 刘文强, 熊海波, 董铮, 薛丹, 盛新年, 赵文锦, 魏秀彩, 李小湘. 水稻蜡质稀少突变体wax1的鉴定及基因定位[J]. 中国水稻科学, 2020, 34(1): 1-7.

Figures/Tables 6

References 29

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引物名称 Primer usage	正向引物(5'→3') Forward primer (5'→3')	反向引物(5'→3') Reverse primer (5'→3')
RM5806	GAATGCTAATTGCGGTTGAAGC	GGATCTTTCCTCCCAATCTTTGC
ID09139	GAAACGGAGGGAGTATATCT	CTGTTTCCGATTCTAAGAGC
RM25414	GCTACCCAACCGCCTCCATAGC	GATCGATCGGAGAGCTTGAGTTGG
ID09071	AAGGTATTTCCCTCTCCTGA	AAGAAGAAGAAAAACCCAGG
P1	TCAACGAACACCAACAAGC	CAGATTATCGGCGGTCACT
OsActin	CATTGGTGCTGAGCGTTTCC	AGAAACAAGCAGGAGGACGG
OSH6	GGTATTTGGTGGTGATTGTGTC	AACCCTTGAGTTGAGCCATT
OsIAA1	ACCAAGAGCCGCTCAATGAG	ATCACACGTGGGCGAACATC
OsIAA4	GCTCTTGCTGGATGGGTATGA	AGGTGATGGGCGTCTTGAAC
OsIAA6	GGCTATCGTCAGCTGTCAAACA	GCAATTTGCGCATTAGTTTGG
OsIAA9	CGAGAAGAAAATGGCCAATGA	ATCCCCATCACCATCCTCGTA
OsIAA15	CGTCCCCTGGAAAATGTTTG	TGCTAAATTGACTGCTTCAGAGCTT
OsIAA18	AAGAATGTGGGAAGGAGCTAACG	ATGGTGGTGAGGGACAGCAT

引物名称 Primer usage	正向引物(5'→3') Forward primer (5'→3')	反向引物(5'→3') Reverse primer (5'→3')
RM5806	GAATGCTAATTGCGGTTGAAGC	GGATCTTTCCTCCCAATCTTTGC
ID09139	GAAACGGAGGGAGTATATCT	CTGTTTCCGATTCTAAGAGC
RM25414	GCTACCCAACCGCCTCCATAGC	GATCGATCGGAGAGCTTGAGTTGG
ID09071	AAGGTATTTCCCTCTCCTGA	AAGAAGAAGAAAAACCCAGG
P1	TCAACGAACACCAACAAGC	CAGATTATCGGCGGTCACT
OsActin	CATTGGTGCTGAGCGTTTCC	AGAAACAAGCAGGAGGACGG
OSH6	GGTATTTGGTGGTGATTGTGTC	AACCCTTGAGTTGAGCCATT
OsIAA1	ACCAAGAGCCGCTCAATGAG	ATCACACGTGGGCGAACATC
OsIAA4	GCTCTTGCTGGATGGGTATGA	AGGTGATGGGCGTCTTGAAC
OsIAA6	GGCTATCGTCAGCTGTCAAACA	GCAATTTGCGCATTAGTTTGG
OsIAA9	CGAGAAGAAAATGGCCAATGA	ATCCCCATCACCATCCTCGTA
OsIAA15	CGTCCCCTGGAAAATGTTTG	TGCTAAATTGACTGCTTCAGAGCTT
OsIAA18	AAGAATGTGGGAAGGAGCTAACG	ATGGTGGTGAGGGACAGCAT

材料 Material	株高 Plant height / cm	有效穗数 No. of effective panicles	每穗总粒数 No. of spikelets per panicle	结实率 Seed-setting rate /%	千粒重 1000-grain weight /g
WT	81.4±3.4	5.5±0.5	134.3±6.7	81.8±6.3	23.9±0.4
wax1	61.2±2.3^**	7.4±0.2^**	94.6±4.0^**	77.7±2.5	21.9±0.3^**

材料 Material	株高 Plant height / cm	有效穗数 No. of effective panicles	每穗总粒数 No. of spikelets per panicle	结实率 Seed-setting rate /%	千粒重 1000-grain weight /g
WT	81.4±3.4	5.5±0.5	134.3±6.7	81.8±6.3	23.9±0.4
wax1	61.2±2.3^**	7.4±0.2^**	94.6±4.0^**	77.7±2.5	21.9±0.3^**