水稻短宽粒基因SWG1的图位克隆

doi:10.3864/j.issn.0578-1752.2023.07.005

摘要/Abstract

摘要：

【目的】水稻产量由单位面积有效穗数、每穗粒数和粒重3个因素构成，其中，粒重主要由水稻的籽粒形态决定。筛选和鉴定新的粒型突变材料和基因，可为产量性状的分子设计育种奠定基础。【方法】在籼稻保持系西大1B（XD1B）的甲基磺酸乙酯（EMS）诱变群体中鉴定到一个短宽粒突变体short and widen grain 1（swg1）；分析籽粒形态和其他农艺性状，并对颖壳进行组织细胞学观察分析；运用BSA法进行基因定位；通过遗传互补试验确定候选基因；采用qRT-PCR分析该基因的表达模式及其他粒型相关基因和细胞发育基因的表达水平。【结果】农艺性状分析发现，与野生型相比，swg1突变体粒长显著降低，粒宽显著增加，表现出短宽粒的表型；进一步组织和细胞学分析，发现突变体颖壳纵向细胞变短是粒长变短的主要原因，而粒宽增加是由于颖壳横向细胞数目和细胞大小同时增加。遗传分析结果表明，该突变性状受隐性单基因控制，通过图位克隆与遗传互补验证，确定候选基因为LOC_Os07g42410，编码一个植物特异转录因子。qRT-PCR分析发现该基因表达无明显的组织特异性，在茎、叶、幼穗中表达强烈。通过对已知粒型相关基因、细胞周期和细胞扩展相关基因进行分析，发现通过正向调控颖壳横向细胞数目和（或）细胞大小决定粒宽的GS5和GW8在突变体中上调明显，而正向调控纵向细胞数目和大小并负向调控横向细胞数目和大小的GW7/GL7在突变体明显下调；另外，部分与细胞周期和细胞扩展相关的基因也在突变体和野生型之间的表达也呈现显著差异。【结论】 SWG1编码一个植物特异的转录因子，通过调控粒型基因（GS5、GW8和GW7/GL7等）影响颖壳的细胞增殖和细胞扩展，从而决定水稻籽粒长度和宽度。

关键词: 水稻, 籽粒形态, 颖壳发育, 图位克隆

Abstract:

【Objective】 Rice yield is composed of effective panicle number per unit area, grains per panicle and grain weight, in which grain weight is mainly determined by grain morphology. Screening and identification of new grain type mutation materials and genes can lay a foundation for molecular design breeding of yield traits. 【Method】 A short and wide grain mutant short and widen grain1 (swg1) was identified in the mutant population of indica rice maintainer line Xida1B(XD1B) induced by ethyl methane sulfonate (EMS). The grain morphology and other agronomic characters were analyzed, and the glume was observed and analyzed by histocytology. Gene mapping was carried out by BSA method, and candidate genes were identified by genetic complementarity experiment. qRT-PCR was used to analyze the expression pattern of the gene and the expression level of other genes related to grain shape and cell development.【Result】 The analysis of agronomic characters showed that the grain length of swg1 mutant was significantly lower and the grain width was significantly higher than that of wild type, showing the phenotype of short and wide grains, and further histological and cytological analysis showed that the shortening of longitudinal cells of glume was the main reason for the shortening of grain length, while the increase of grain width was due to the increase of the number and size of transverse cells of glume at the same time. The results of genetic analysis showed that the mutation was controlled by a recessive single gene, and the candidate gene for SWG1 was determined to be LOC_Os07g42410 by map-based cloning and genetic complementary verification, which encoded a plant-specific transcription factor. qRT-PCR analysis showed that the expression of this gene had no obvious tissue specificity, and its expression is strong in stem, leaf and young panicle. According to the analysis of the expression of known genes related to grain shape, cell cycle and cell expansion, it was found that GS5 and GW8, which positively regulate the number and/or size of glume transverse cells to determine grain width, were significantly up-regulated in the mutants, while GW7/GL7 genes, which positively regulated the number and size of longitudinal cells and negatively regulated the number and size of transverse cells, were significantly down-regulated in the mutants. Some genes related to cell cycle and cell expansion also showed significant differences between mutants and wild types. 【Conclusion】 SWG1 encodes a plant-specific transcription factor, which affects glume cell proliferation and cell expansion by regulating grain shape genes GS5, GW8 and GW7/GL7, thus determining rice grain length and width.

Key words: rice (Oryza sativa L.), grain, development of glumes, gene mapping

朱洪慧, 李映姿, 高远卓, 林泓, 王成洋, 晏紫仪, 彭瀚平, 李田野, 熊茂, 李云峰. 水稻短宽粒基因SWG1的图位克隆[J]. 中国农业科学, 2023, 56(7): 1260-1274.

ZHU HongHui, LI YingZi, GAO YuanZhuo, LIN Hong, WANG ChengYang, YAN ZiYi, PENG HanPing, LI TianYe, XIONG Mao, LI YunFeng. Map-Based Cloning of the SHORT AND WIDEN GRAIN 1 Gene in Rice (Oryza sativa L.)[J]. Scientia Agricultura Sinica, 2023, 56(7): 1260-1274.

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引物名称 Primer name	正向序列 Forward sequence (5′-3′)	反向序列 Reverse sequences (5′-3′)	用途 Purpose
RM234	TTCAGCCAAGAACAGAACAGTGG	CTTCTCTTCATCCTCCTCCTTGG	定位 Mapping
RM21934	GCAGCAGTGTTGTACTGTTCTTCG	GGGAGGGCTTACTCTGTATCAGG
RM21968	GTCATCGACATCCAGCTTCAAGG	TCGACTCCATTATCGGATGTGC
RM21979	ACGCCGCGAGATAGTTATCAACC	GAGCACGGTGTTGTAGGAGACG
ACTIN	TGGCATCTCTCAGCACATTCC	TGCACAATGGATGGGTCAGA	qRT-PCR
qSWG1	TGCGTGATAGCCTAGAACGAAG	CTGGAATCAGCACTCCTGGATG
qOsEXPB2	TCGTCTACACCAACGACTGG	CATGAACGGGTACTGGTTGG
qOsEXPB3	TTCTCGTCGATGACCTCCTG	AGGGTGGTTGACGCATCTTA
qOsEXPB4	GTCGGTCTGTGTTGCGATTTG	CCTCCATTTCCCACACAGCTT
qOsEXPB5	AAGGCTGTGGCTTGATTGACA	TTAGGCCCAATTTTGCTATTTTG
qOsEXPB7	ACGGTGATCATCACGGACAT	TCGAAGTGGTACAGCGACACT
qOsEXPB10	TGACCAACTACAACGTGGTCCC	GCCAGTGTATGTTTTGCCGAAG
qCycA1;1	CTTTCGGTTGACGAGACGATGT	CGCTGCAAGGAACCTAGAACTG
qCycA2;1	AATATTGAGCGAAACAGGGACAG	AGGAAGCACACATTTGAGGATTT
qCycA3;2	AGGTTGTCAAGATGGAGAGCGA	CGCTTTTTGTCTTCCTGGCA
qCycB1;1	CACTCTCAAGCACCACACTGGA	ACAACCCTCAGCTTGCTCTCAG
qCycB2;1	CGCTTGCAGCAACCGAGTA	CATCCATCAGCTCAAACTTGTGAT
qCycB2;2	CTCAAGGCTGCACAATCTGACA	GCATTGACGGCTGGAATTTG
qGS5	CATTCCATGCAAATGCCAGTGGAC	CAGCCCTGCTTTGATGAGCTTG
qGW2	CAGCAGCGCATTCCCAGTTTTC	GTGGTCAGCCGAGCACTCTC
qGL3.1	TCACAACTCCCAGGATAGG	TTTGTCTCGCTCGCTCAT
qGW8	AGGAGTTTGATGAGGCCAAG	GCGTGTAGTATGGGCTCTCC
qGL6	TCTGCTTGACATGTGACAGGA	GAGGATGTTGGAGAGGTCGC
qGL7	CCCCTAGCATCGACACCAAG	CGGGTTCCAGCACTCCTCT
qTGW2	CCCGGAAACTGTCTCATCAC	GAACTGAGCCCTCATCTTGG
qTGW6	TTGAACTTGCAAAAGGCAGA	CGGTTCCCCTAATGCAGAT
qGS2	ACCACTCTCTTTACCCTGCT	CGGGTTCCAGCACTCCTCT
qGS3	CATCGGAGAAGCGAAGTCAT	TTGAGGTTGAAGGAGGAGGA

序号Number	基因名称 Locus name	基因注释 Gene annotation
1	LOC_Os07g42390	含有DUF581结构域的蛋白，表达DUF581 domain containing protein, expressed
2	LOC_Os07g42395	DNA定向RNA聚合酶II亚基RPB9，推测，表达 DNA-directed RNA polymerase II subunit RPB9, putative, expressed
3	LOC_Os07g42400	转座子蛋白，推测，未分类，表达Transposon protein, putative, unclassified, expressed
4	LOC_Os07g42410	转录因子Transcription factor
5	LOC_Os07g42420	3-氧酰基合酶，推测，表达3-oxoacyl-synthase, putative, expressed
6	LOC_Os07g42430	表达蛋白质Expressed protein
7	LOC_Os07g42440	乙醇酸氧化酶Glycolate oxidase
8	LOC_Os07g42450	核糖体蛋白S2，推测，表达Ribosomal protein S2, putative, expressed
9	LOC_Os07g42460	转座子蛋白，推测，未分类，表达Transposon protein, putative, unclassified, expressed
10	LOC_Os07g42470	表达蛋白质Expressed protein
11	LOC_Os07g42490	水稻蔗糖合酶Sucrose synthase in rice
12	LOC_Os07g42500	FYVE锌指结构域蛋白，表达FYVE zinc finger domain containing protein, expressed
13	LOC_Os07g42510	含有AP2结构域的蛋白，表达AP2 domain containing protein, expressed
14	LOC_Os07g42520	客观的，推测，表达Dirigent, putative, expressed
15	LOC_Os07g42530	转座子蛋白，推测，未分类，表达Transposon protein, putative, unclassified, expressed
16	LOC_Os07g42540	转座子蛋白，推测，Pong亚类Transposon protein, putative, Pong sub-class
17	LOC_Os07g42550	表达蛋白质Expressed protein
18	LOC_Os07g42560	表达蛋白质Expressed protein