Identification of a New OsBRI1 Weak Allele and Analysis of its Function in Grain Size Control

doi:10.11983/CBB19239

Abstract

Abstract: Rice (Oryza sativa) grain size and grain weight are key agronomic traits that affect rice yield. Cloning and study of grain size genes are helpful to increase rice production. In order to further understand the mechanism of rice grain size control, a set of mutants with altered grain size from an EMS-treated elite japonica cultivar KYJ (Kuanyejing) were isolated. smg12 exhibits small grains, short plants, and reduced number of primary branches and secondary branches. Genetic analyses show that the smg12 mutant phenotypes are controlled by a single recessive gene. Our celluar analyses show that the small grain size phenotype of smg12 is caused by the decrease in cell size of glumes, indicating that SMG12 affects cell expansion. By using the Mutmap method, we reveal that the candidate gene for SMG12 is OsBRI1, which encodes a brassinolide receptor kinase. The smg12 mutant causes a substitution of the 2 074^th base (C to T) in OsBRI1, which results in an amino acid change (proline to serine). Therefore, this study identified a new mutant allele of OsBRI1 and provides a cellular and molecular basis for BR-mediated grain size control in rice.

Key words: rice, grain size, cell expansion, Mutmap, OsBRI1

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[J]. Chinese Bulletin of Botany, 2020, 55(3): 279-286.

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URL: https://www.chinbullbotany.com/EN/10.11983/CBB19239

https://www.chinbullbotany.com/EN/Y2020/V55/I3/279

Figures/Tables 6

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.

Table 1 Primer and enzyme for dCAPS analysis

Primer name	Primer sequence (5'→3')	DNA fragment length (bp)	Enzyme
smg12-1	CTTTCTCGGCACTTTCCTTG	154	HphI
smg12-1	CTATGGTCACATGGTGGCGGTG	154	HphI

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).

Table 2 Genetic analysis of the rice mutant smg12

Hybrid combinations	Phenotype of F₁	F₂ generation			χ²_3:1
Hybrid combinations	Phenotype of F₁	Wild-phenotypic plant number	Mutant-phenotypic plant number	Total number	χ²_3:1
smg12/KYJ	Wild type	168	52	220	0.151

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	/

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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