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Acta Agronomica Sinica ›› 2019, Vol. 45 ›› Issue (6): 807-817.doi: 10.3724/SP.J.1006.2019.81090

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

Fine mapping and candidate gene analysis of awn inhibiting gene B2 in common wheat

Di JIN1,*,Dong-Zhi WANG2,*,Huan-Xue WANG3,Run-Zhi LI3,Shu-Lin CHEN1,Wen-Long YANG2,Ai-Min ZHANG2,Dong-Cheng LIU2,4,*(),Ke-Hui ZHAN1,*()   

  1. 1 Agronomy College of Henan Agricultural University, Zhengzhou 450002, Henan, China
    2 State Key Laboratory of Plant Cell and Chromosome Engineering / Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China
    3 College of Plant Science and Technology, Beijing University of Agriculture, Beijing 102206, China
    4 Biology and Agriculture Research Center, University of Science and Technology Beijing, Beijing 100024, China
  • Received:2018-12-19 Accepted:2019-01-19 Online:2019-06-12 Published:2019-06-12
  • Contact: Di JIN,Dong-Zhi WANG,Dong-Cheng LIU,Ke-Hui ZHAN E-mail:dongchengliu@ustb.edu.cn;kh486@163.com
  • Supported by:
    This study was supported by the National Key Research and Development Program of China(2016YFD0101800);the National Natural Science Foundation of China(31571643)

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Abstract:

Awn is one of the important photosynthetic organs in common wheat and plays a vital role in yield potential and environmental adaptation. At present, the inheritance and development of wheat awn have been not systematically studied, and cloning or fine mapping of related genes are seldom reported. In this study, the genetics and candidate genes conferring the short awn of a Chinese wheat landrace ‘Liuzhutou’ were investigated. Longitudinal section of awn showed cell size of short awn was much shorter than that of long awn. Using Wheat660K SNP chip based bulked segregant analysis (BSA) and SL-F2 population derived from ‘Liuzhutou’ (short awn) and modern cultivar ‘Shiai 1’ (long awn), the awn inhibiting gene in ‘Liuzhutou’ was mapped in a 4.84 Mb interval on chromosome 6BL and predicted as previously characterized B2. A good micro-collinearity of B2 region was observed among chromosomes 6B, 6A, 6D of Chinese Spring and chromosome 6B of AK58, there were 61 genes annotated in the 4.84 Mb B2 region, five of which were specifically expressed in the developing spike of Chinese Spring, and TraesCS6B02G264400 was differentially expressed between Chinese Spring and Azhurnaya. These data provide important clues for cloning the B2 gene, dissecting the developing mechanism of wheat awn and its application in molecular breeding.

Key words: common wheat, awn, B2, fine mapping, BSA, Wheat660K

Table 1

Primers used in this study"

引物名称
Primer ID		 正向引物序列
Forward sequence (5'-3')		 反向引物序列
Reverse sequence (5'-3')		 内切酶
Restriction enzyme		 片段大小
Size (bp)
B2精细定位 Markers for mapping B2
Xgwm88 * CACTACAACTATGCGCTCGC TCCATTGGCTTCTCTCTCAA — 120
SSR43 ACTATCACAAGCCGGAGTAA AGTGGTTAAATCGCCACTAA — 172
SSR122 CATGTTACCACCAAAGGATT CTATGTGGCTGACCGTTACT — 205
EST-499300LC TCCAAAGAATCCTGGAGGTG TGTTCTGACCATCCCACGTA Pvu II 393/270
dCAPS-18 GACCACCCGACACCGCCGCTGCA GCTCATCTCCAATAGCTCGC Pst I 204/184
dCAPS-19 CAAAACTTCTTGGGAGAATATAC CCCTGACATCGTCTCGAACT Mlu I 192/172
dCAPS-4 GTAAGCAGCGAGGTTAGCACTGC CTGGTGCTCATGAGTTGTGG Pst I 296/276
EST-263300 CGTGTGCTTAAATGCTCCATGG GCTGACTTATGCTGCTTTACTTTCTAA Taq I 521/472
CAPS-12 ATTGATGCAAAGGAACCAGG GAGTTCCATGGCCTCATTGT Eco RI 308/236
CAPS-14 GGAAGGTCCACTTTGCATGT CACCACAGGGACGAAATTCT Bam HI 281/265
dCAPS-39 GTTTTCTTTGTAGAAGTCCCATA AAACACCTGAAAATCGTGGC Nde I 185/165
EST-265400 CCATCAGTCCAACCATGACTTGT TAACATAAGCGTCGCTGTGGC Pst I 246/(111+135)
CAPS-19 TGTCCTTGAAGGGGGTAGTG TCCTGCATTATGCCACAAAA Nde I 483/(217+266)
WABM232658 * AAGTTCGCCTCTTCACCAGT TCTGCCCCTACATCTGTTGC — 130
SSR74 CATAATCACAATTCATCGGA TTCATACCTGACCCATCTTC — 214
SSR116 AGAATCAGTTTTCAGCCAGA AACTATCCCGTATACTTGCG — 168
WABM214868 * GGGTGCCTGAACATTGATGC CCCCAAGTGCTGTCGTGTAT — 126
候选基因克隆 Gene cloning primers
6B01G262600 AGCGAAGCAGTCAGTACTCAGT CTTGCATGGATAATCGAAACATAGGA — 1041
6B01G263300 CGTGTGCTTAAATGCTCCATGG GCTGACTTATGCTGCTTTACTTTCTAA — 1110
6B01G264600 GCCCATTTTTGCAGTCATGACATCA CGGGATGAAAACCTGCTCTTC — 1760
6B01G265400 CCATCAGTCCAACCATGACTTGT TAACATAAGCGTCGCTGTGGC — 1071
6B01G504500LC CACACACACACACACCAAAACTC GGCATACTGCGAACAACCCA — 985
6B01G505500LC AGCAGCTGGGTGAAGTTAACTA TTCCAGCCCCCTTTTACAAGAT — 1437

Fig. 1

Longitudinal section of middle region in long awn (A) and short awn (B)"

Fig. 2

Population construction and analysis of Wheat660K SNP chip (A) Population construction of SL-F2 and the awn character of ‘ShiAi 1’, ‘Liuzhutou’ F1 and F2; (B) The distribution of differential SNP locus on each chromosome and between bulks of SL-F2 population; (C) The distribution of differential SNP locus on chromosome 6B between bulks of SL-F2 population. Un: unknown."

Table 2

Chi-square test and genetic analysis of SL-F2 population"

单株数
No. of F2 plants		 观察值 Observed value 理论值 Expected value 理论比值
Expected ratio		 χ2值
χ2-value		 P值
P-value
长芒
Long awn		 短芒
Short awn		 长芒
Long awn		 短芒
Short awn
1413 350 1063 353 1060 3:1 0.04 0.84

Fig. 3

Fine mapping of awn inhibiting gene B2 (A) The linkage map of B2 locus in SL-F2 population; (B) The physical map of B2 locus in SL-F2 population; (C) The marker genotype and progeny validation phenotype of key recombinants in SL-F2 population."

Fig. 4

Micro-collinearity analysis of B2 region From left to right are chromosome 6B of AK58, chromosomes 6B, 6A, and 6D of CS; genes in red and cyan colors indicate no homologous genes on chromosomes 6B of CS and AK58, respectively."

Fig. 5

Annotation and expression profiling of genes in B2 region The expression level of each sample is printed as deep blue representing the lowest value to deep red representing the highest value in the heat map. The gene names are followed closely by their annotation information, and the genes in red indicate that its expression reaches the threshold of Fold-Change (TPMSpike/TPMMean (Leaf, Root, Grain)) ≥ 2 in Chinese Spring."

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