Scientia Agricultura Sinica ›› 2020, Vol. 53 ›› Issue (16): 3205-3213.doi: 10.3864/j.issn.0578-1752.2020.16.001

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Genome-Wide Association Study and Candidate Gene Mining of Tillering Number in Japonica Rice

ZHANG JiFeng(),LIU HuaDong,WANG JingGuo,LIU HuaLong,SUN Jian,YANG LuoMiao,JIA Yan,WU WenShen,ZHENG HongLiang(),ZOU DeTang()   

  1. Key Laboratory of Germplasm Enhancement, Physiology and Ecology of Food Crops in Cold Region, Ministry of Education, Northeast Agricultural University, Harbin 150030
  • Received:2019-10-18 Accepted:2020-01-13 Online:2020-08-16 Published:2020-08-27
  • Contact: HongLiang ZHENG,DeTang ZOU E-mail:13804689362@163.com;zhenghongliang008@126.com;zoudtneau@126.com

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Abstract:

【Objective】Genome-wide association study (GWAS) was used to detect SNP loci that were significantly related to tillering number in japonica rice, and to screen candidate genes affecting tillering number 【Method】This study used 295 japonica rice varieties from around the world, in 2018 and 2019 survey from the tillering stage of rice tillering number, the combination of high throughput sequencing weight gain high quality 788396 polymorphism, SNP, using TASSEL 5.0 software genome-wide association analysis of MLM model, using the GEC software to calculate the number of effective independent SNPS for the determination of threshold value, determine the significance of SNP markers associated with target traits. Based on the peak SNP detected in two years and the LD attenuation distance of each chromosome in rice, the major QTLs for co-localization of biennial number were determined, and the non-synonymous SNPs and promoter regions of all gene exon regions in the QTL interval were further extracted. SNPs were subjected to haplotype analysis, and then combined with gene annotation to screen candidate genes affecting japonica rice tillering number. 【Result】 The tillering number of 295 japonica rice varieties were basically the same in 2018 and 2019, and they all had a large phenotypic distribution. Under the threshold of P < 5.46 × 10-6, three QTLs (qTiller8, qTiller9 and qTiller10) related to tillering number of Japonica rice were identified on chromosomes 8, 9 and 10 by genome-wide association analysis. qTiller9 jointly detected that the contribution rate of phenotype in 2018 and 2019 was 11.86% and 10.61%, respectively. qtiller8 and qtiller10 were only detected in 2018, and the contribution rate of phenotype was 10.61% The rates were 9.36% and 9.10%, respectively. Haplotype analysis of all 15 genes in qTiller9 interval showed that there were 6 genes in qTiller9 interval (LOC_Os09g25090, LOC_Os09g25100, LOC_Os09g25150, LOC_Os09g25190, LOC_Os09g25200 and LOC_Os09g25220). LOC_Os09g25090 was divided into two haplotypes by promoter SNP, and the tillering number of hap2 (TAA) was significantly higher than that of hap1 (AGG). LOC_Os09g25100 was divided into two haplotypes by non-synonymy mutation SNP, and hap2 (GAGA) had a significantly higher tillering number than hap1 (AGCC). LOC_Os09g25150 was divided into two haplotypes by non-synonymy mutation SNP, and the tillering number of hap2 (ATG) was significantly higher than that of hap1 (GCC). LOC_Os09g25190 was divided into two haplotypes by promoter SNP and the tillering number of hap2 (GCATCGCATCGACGCCGA) was significantly higher than that of hap1 (ATGCTGATGAAGTCATCC). LOC_Os09g25200 was divided into two haplotypes by non-synonymic mutant SNP and the tillering number of hap2 (TAG) was significantly higher than that of hap1 (AGA). LOC_Os09g25220 was divided into two haplotypes by non-synonymic mutant SNP, and the tillering number of hap1 (GG) was significantly higher than that of hap2 (AA).Combined with gene annotation, it was found that LOC_Os09g25090 and LOC_Os09g25100 both predicted the encoding of calcineurin dependent protein kinases and were Ca2+ sensors necessary for abscisic acid (ABA) expression. Previous studies have shown that ABA can affect both tillering number and branch number of Arabidopsis. Therefore, LOC_Os09g25090 and LOC_Os09g25100 are candidate genes affecting the tillering number in japonica rice. 【Conclusion】LOC_Os09g25090 and LOC_Os09g25100 were screened as candidate genes affecting the tiller number of japonica rice.

Key words: japonica rice, tillering number, genome-wide association study, haplotype analysis, candidate gene

Table 1

The phenotypic values of tillering number among 295 japonica rice varieties in two years"

年份 Year 均值±标准差 Mean±standard 变幅 Range 变异系数 CV (%) 偏度 Skewness 峰度 Kurtosis
2018 18.35±5.98 7.00—49.00 32.58 1.31 0.49
2019 15.75±4.65 6.00—33.80 29.55 3.09 0.05

Fig. 1

Histogram of tillering number frequency distribution under 2-year environmental conditions"

Table 2

Significant loci associated with tillering number in japonica rice"

QTL 年份
Year		 峰值SNP
Peak SNP		 染色体
Chr.		 P值
P value		 表型贡献率
R2(%)		 已定位QTL
QTL located
qTiller8 2018 7527329 8 2.02E-06 9.36
qTiller9 2018 15065450 9 6.99E-08 11.86 qNOT9-1[29]
qTiller10 2018 15406671 10 2.64E-06 9.10
qTiller9 2019 15060619 9 2.55E-07 10.61 qNOT9-1[29]

Fig. 2

Manhattan plots and quantile-quantile (Q-Q) plots of genome-wide association studies for the tillering number A and B indicate the result of 2018 and 2019, respective"

Table 3

Candidate gene haplotype group and the composition of each haplotype SNP"

基因
Gene		 单倍型1/品种数
hap1/Number		 单倍型2/品种数
hap2/Number		 单倍型3/品种数
hap3/Number
LOC_Os09g25090 AGG/182 TAA/81
LOC_Os09g25100 AGCC/188 GAGA/89
LOC_Os09g25150 ATG/190 GCC/69 GTC/17
LOC_Os09g25190 ATGCTGATGAAGTCATCC/126 GCATCGCATCGACGCCGA/76
LOC_Os09g25200 AGA/181 TAG/89
LOC_Os09g25220 GG/97 AA/187

Fig. 3

Boxplots for the tillering number based on the haplotypes(hap) for candidate gene Green, yellow and blue indicate phenotypic result for hap1, hap2 and hap3, respectively"

Table 4

Candidate gene of gene annotation"

基因 Gene 基因功能注释 Gene annotation
LOC_Os09g25090 钙调蛋白依赖性蛋白激酶
Calmodulin depedent protein kinases
LOC_Os09g25100 钙调蛋白依赖性蛋白激酶
Calmodulin depedent protein kinases
LOC_Os09g25150 肉桂酰辅酶A还原酶
Cinnamoyl-CoA reductase
LOC_Os09g25190 蛋白结合蛋白Protein binding protein
LOC_Os09g25200 蛋白结合蛋白Protein binding protein
LOC_Os09g25220 蛋白结合蛋白Protein binding protein
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