植物学报 ›› 2020, Vol. 55 ›› Issue (3): 279-286.DOI: 10.11983/CBB19239
管柳蓉1,2,刘祖培2,3,徐冉2,段朋根2,张国政2,3,于海跃2,3,李静2,3,罗越华1,*(),李云海2,*(
)
收稿日期:
2019-12-25
接受日期:
2020-03-23
出版日期:
2020-05-01
发布日期:
2020-07-06
通讯作者:
罗越华,李云海
基金资助:
Liurong Guan1,2,Zupei Liu2,3,Ran Xu2,Penggen Duan2,Guozheng Zhang2,3,Haiyue Yu2,3,Jing Li2,3,Yuehua Luo1,*(),Yunhai Li2,*(
)
Received:
2019-12-25
Accepted:
2020-03-23
Online:
2020-05-01
Published:
2020-07-06
Contact:
Yuehua Luo,Yunhai Li
摘要:
水稻(Oryza sativa)籽粒大小是影响其产量的关键农艺性状, 克隆并研究水稻籽粒大小相关基因对于提高水稻产量具有重要意义。为深入探究水稻籽粒大小的调控机制, 通过EMS诱变品种宽叶粳(KYJ), 分离了一系列水稻籽粒大小改变的突变体, 其中smg12表现为籽粒变小, 株高变矮, 一级枝梗数和二级枝梗数减少。遗传分析表明, 该小粒突变体受隐性单基因控制。细胞学分析显示, 该突变体颖壳纵向细胞长度显著变短, 表明SMG12主要影响细胞扩展。利用Mutmap方法对候选基因进行克隆, 筛选出SMG12的候选基因OsBRI1, 该基因编码油菜素内酯受体激酶。OsBRI1外显子上的第2 074个碱基发生了由C到T的置换, 产生非同义突变, 使得该位置编码的脯氨酸变为丝氨酸, 从而影响OsBRI1的功能。综上, 该研究鉴定了OsBRI1基因的1个新等位变异, 揭示了油菜素内酯途径调控水稻籽粒大小的细胞和分子基础。
管柳蓉, 刘祖培, 徐冉, 段朋根, 张国政, 于海跃, 李静, 罗越华, 李云海. 一个新的OsBRI1弱等位突变体的鉴定及其调控种子大小的功能研究. 植物学报, 2020, 55(3): 279-286.
Liurong Guan, Zupei Liu, Ran Xu, Penggen Duan, Guozheng Zhang, Haiyue Yu, Jing Li, Yuehua Luo, Yunhai Li. Identification of a New OsBRI1 Weak Allele and Analysis of its Function in Grain Size Control. Chinese Bulletin of Botany, 2020, 55(3): 279-286.
图1 水稻突变体smg12的表型分析 (A) KYJ (野生型)和smg12突变体表型(Bar=10 cm); (B) KYJ和smg12的籽粒(Bars=1 cm); (C) 粒宽; (D) 粒长; (E) 一级枝梗数; (F) 二级枝梗数; (G) 株高; (H) 千粒重。显著性分析采用t检验, * 表示差异显著(P<0.05), ** 表示差异极显著(P<0.01)。
Figure 1 Analysis of rice smg12 mutant phenotypes (A) KYJ (wild type) and smg12 mutant phenotype (Bar=10 cm); (B) Grains of KYJ and smg12 (Bars=1 cm); (C) Grain width; (D) Grain length; (E) Number of primary branches; (F) Number of secondary branches; (G) Plant height; (H) Thousand seed weight. Significance is determined using t-test, * indicates significant differences at P<0.05, ** indicates significant differences at P<0.01.
Primer name | Primer sequence (5'→3') | DNA fragment length (bp) | Enzyme |
---|---|---|---|
smg12-1 | CTTTCTCGGCACTTTCCTTG | 154 | HphI |
CTATGGTCACATGGTGGCGGTG |
表1 用于dCAPS分析的引物和酶
Table 1 Primer and enzyme for dCAPS analysis
Primer name | Primer sequence (5'→3') | DNA fragment length (bp) | Enzyme |
---|---|---|---|
smg12-1 | CTTTCTCGGCACTTTCCTTG | 154 | HphI |
CTATGGTCACATGGTGGCGGTG |
图2 SMG12调控水稻颖壳细胞大小 (A) 颖壳纵向细胞数目; (B) 颖壳横向细胞数目; (C) 颖壳单个细胞纵向长度; (D) 颖壳单个细胞横向宽度; (E) 细胞学分析图(Bars=0.1 mm)。** 表示突变体与野生型之间差异极显著(P< 0.01)。
Figure 2 SMG12 regulates cell size in grain hulls of rice (A) Outer epidermal cell number in the grain-length direction of the glume; (B) Outer epidermal cell number in the grain-width direction of the glume; (C) The longitudinal length of a single cell in the glume; (D) Transverse width of a single cell in the glume; (E) Cytological analysis diagram (Bars= 0.1 mm). ** indicates extremely significant difference between the mutant and the wild type (P<0.01).
Hybrid combinations | Phenotype of F1 | F2 generation | χ23:1 | ||
---|---|---|---|---|---|
Wild-phenotypic plant number | Mutant-phenotypic plant number | Total number | |||
smg12/KYJ | Wild type | 168 | 52 | 220 | 0.151 |
表2 水稻突变体smg12的遗传分析
Table 2 Genetic analysis of the rice mutant smg12
Hybrid combinations | Phenotype of F1 | F2 generation | χ23:1 | ||
---|---|---|---|---|---|
Wild-phenotypic plant number | Mutant-phenotypic plant number | Total number | |||
smg12/KYJ | Wild type | 168 | 52 | 220 | 0.151 |
Chromosomal position | Physical location (bp) | Genotype (KYJ/smg12) | Gene location | Gene locus name | Frequency of sequencing (KYJ/smg12) | Mutation type |
---|---|---|---|---|---|---|
Chr. 1 | 15686971 | G/A | Upstream of the gene | LOC_Os01g28040 | 0/6 | / |
Chr. 1 | 27750517 | G/A | In the gene compartment | LOC_Os01g48420; LOC_Os01g48430 | 0/12 | / |
Chr. 1 | 28534819 | G/A | Upstream and downstream of the gene | LOC_Os01g49630; LOC_Os01g49614 | 0/14 | / |
Chr. 1 | 29397508 | G/A | In the gene compartment | LOC_Os01g51140; LOC_Os01g51154 | 0/12 | / |
Chr. 1 | 29788896 | G/A | Upstream of the gene | LOC_Os01g51810 | 0/17 | / |
Chr. 1 | 29929259 | G/A | Exon | LOC_Os01g52050 | 0/19 | Changes in amino acids |
Chr. 1 | 33892012 | T/A | Upstream of the gene | LOC_Os01g58620 | 0/1 | / |
Chr. 1 | 39503252 | G/A | Downstream of the gene | LOC_Os01g67980 | 0/11 | / |
Chr. 1 | 30392222 | G/A | Upstream of the gene | LOC_Os01g52840; LOC_Os01g52851 | 0/5 | / |
Chr. 1 | 33745383 | G/A | Intron | LOC_Os01g58400 | 0/8 | / |
Chr. 3 | 19627588 | C/A | Upstream of the gene | LOC_Os03g35390 | 0/6 | / |
Chr. 6 | 17710551 | G/A | Intron | LOC_Os06g30610 | 0/8 | / |
表3 smg12突变体的候选SNPs分析
Table 3 Analysis of the candidate SNPs for the smg12 mutant
Chromosomal position | Physical location (bp) | Genotype (KYJ/smg12) | Gene location | Gene locus name | Frequency of sequencing (KYJ/smg12) | Mutation type |
---|---|---|---|---|---|---|
Chr. 1 | 15686971 | G/A | Upstream of the gene | LOC_Os01g28040 | 0/6 | / |
Chr. 1 | 27750517 | G/A | In the gene compartment | LOC_Os01g48420; LOC_Os01g48430 | 0/12 | / |
Chr. 1 | 28534819 | G/A | Upstream and downstream of the gene | LOC_Os01g49630; LOC_Os01g49614 | 0/14 | / |
Chr. 1 | 29397508 | G/A | In the gene compartment | LOC_Os01g51140; LOC_Os01g51154 | 0/12 | / |
Chr. 1 | 29788896 | G/A | Upstream of the gene | LOC_Os01g51810 | 0/17 | / |
Chr. 1 | 29929259 | G/A | Exon | LOC_Os01g52050 | 0/19 | Changes in amino acids |
Chr. 1 | 33892012 | T/A | Upstream of the gene | LOC_Os01g58620 | 0/1 | / |
Chr. 1 | 39503252 | G/A | Downstream of the gene | LOC_Os01g67980 | 0/11 | / |
Chr. 1 | 30392222 | G/A | Upstream of the gene | LOC_Os01g52840; LOC_Os01g52851 | 0/5 | / |
Chr. 1 | 33745383 | G/A | Intron | LOC_Os01g58400 | 0/8 | / |
Chr. 3 | 19627588 | C/A | Upstream of the gene | LOC_Os03g35390 | 0/6 | / |
Chr. 6 | 17710551 | G/A | Intron | LOC_Os06g30610 | 0/8 | / |
图3 候选基因的鉴定 (A) smg12突变的dCAPS标记验证; (B) 候选基因LOC_Os01g52050的群体连锁分析; (C) LOC_Os01g52050的基因结构图, 开放的方框表示基因5'和3'端的未翻译区, 封闭的方框表示编码区, 且图中标注起始密码子(ATG)和终止密码子(TAG)以及LOC_Os01g52050中的突变位点(C/T); (D) LOC_Os01g52050的蛋白结构域。P/S表示LOC_Os01g52050编码蛋白结构域中的突变位点。
Figure 3 Identification of candidate genes (A) The dCAPS marker was developed to detect the smg12 mutation; (B) Population linkage analysis of the candidate gene LOC_Os01g52050; (C) The LOC_Os01g52050 gene structure, open boxes show the 5' and 3' untranslated regions, the closed box shows the coding sequence, and the start codon (ATG), the stop codon (TAG) and the LOC_Os01g52050 mutation site (C/T) are indicated; (D) Schematic of the LOC_Os01g52050 protein. P/S indicated the LOC_Os01g52050 mutation site.
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