控制高粱分蘖与主茎株高一致性的基因定位

doi:10.3724/SP.J.1006.2019.84111

作物学报 ›› 2019, Vol. 45 ›› Issue (6): 829-838.doi: 10.3724/SP.J.1006.2019.84111

• 作物遗传育种·种质资源·分子遗传学 • 上一篇下一篇

控制高粱分蘖与主茎株高一致性的基因定位

王瑞^1,^*,凌亮^2,^*,詹鹏杰¹,于纪珍¹,楚建强¹,平俊爱^1,^*(),张福耀^1,^*()

1 山西省农业科学院高粱研究所 / 高粱遗传与种质创新山西省重点实验室, 山西榆次 030600
2 山西省农业科学院食用菌研究所, 山西太原030031

收稿日期:2018-08-21 接受日期:2019-01-19 出版日期:2019-06-12 网络出版日期:2019-06-12
通讯作者: 王瑞,凌亮,平俊爱,张福耀
作者简介:王瑞, E-mail: wangrui989@163.com|凌亮, E-mail: 478781770@qq.com
基金资助:
本研究由山西省农业科学院博士研究基金(YBSJJ1606);国家现代农业产业技术体系建设专项(CARS-06);山西省农业科学院优势课题组(YCX2018D2YS11)项目资助

Mapping of genes confessing same height of tiller and main stem in sorghum

Rui WANG^1,^*,Liang LING^2,^*,Peng-Jie ZHAN¹,Ji-Zhen YU¹,Jian-Qiang CHU¹,Jun-Ai PING^1,^*(),Fu-Yao ZHANG^1,^*()

1 Sorghum Institute, Shanxi Academy of Agricultural Sciences / Key Laboratory of Genetic and Germplasm Innovation in Sorghum for Shanxi Province, Yuci 030600, Shanxi, China
2 Institute of Edible Fungi, Shanxi Academy of Agricultural Sciences, Taiyuan 030031, Shanxi, China

Received:2018-08-21 Accepted:2019-01-19 Published:2019-06-12 Published online:2019-06-12
Contact: Rui WANG,Liang LING,Jun-Ai PING,Fu-Yao ZHANG
Supported by:
This study was supported by the Doctoral Research Fund of Shanxi Academy of Agricultural Sciences(YBSJJ1606);the China Agriculture Research System(CARS-06);the Advantage Research Group of Shanxi Academy of Agricultural Sciences (YCX2018D2YS11).

摘要/Abstract

摘要：

用分蘖与主茎株高一致的高粱品系K35-Y5与分蘖明显高于主茎的高粱恢复系1383杂交, F₁自交获得F₂分离群体, 构建两混池, 采用BSA (bulked segregation analysis)和SLAF (specific length amplified fragment sequencing)技术将高粱分蘖与主茎株高一致基因定位。遗传分析表明, 分蘖与主茎株高一致性状由1对隐性核基因控制。参考已公布高粱基因组设计酶切方案, 构建SLAF文库并测序。对高粱参考基因组序列进行电子酶切预测, 确定限制性内切酶为Rsa I + Hae III, 酶切片段长度为364~414 bp; 测序Q30为91.70%, GC含量为45.79%, 达到测序要求; 与水稻的测序数据相比, 高粱的双端比对效率为93.35%, 酶切效率为90.60%, SLAF建库正常。共获得30.80 M reads, 开发出133,246个SLAF标签, 再通过分析SLAF标签的多态性, 检测到319,428个SNP位点。利用SNP-index法和Euclidean distance法及取两者交集进行关联分析, 最后得到一个关联区域, 位于第9染色体上的54,788,026~56,740,873区间内, 关联区域长度1.95 Mb。分析关联区域内的基因在2个亲本之间SNP, 对这些SNP进行变异的注释, 发现4个非同义突变的SNP。经验证, 这4个SNP位点和分蘖与主茎株高一致性状相关。对应到Sobic.009G197901.1、Sobic.009G213300.1和Sobic.009G221200.1三个基因上, 这些基因可能是与性状直接相关的功能基因。

关键词: 高粱, 分蘖与主茎株高一致, SLAF, SNP

Abstract:

In this study, an F₂ population derived from a cross between two sorghum lines with same height and different heights of tiller and main stem respectively was used to construct pools. In order to map genes related to same height of tiller and main stem, BSA and SLAF-seq technique were developed. Genetic analysis showed that the trait of same height of tiller and main stem was controlled by a single recessive nuclear gene. Reference genome of sorghum was used to design markers by simulating the number of markers produced by different enzymes. The SLAF library was conducted and sequenced by paired-end sequencing. The restriction enzyme was Rsa I + Hae III. The fragment length was 364-414 bp. The quality of Q30 was up to 91.70% and the GC content (45.79%) was low enough to perform sequencing. Compared with the sequencing data of rice, the construction of SLAF library fitted well to the standard, with its paired-end mapped reads reaching to 93.35% and normal digestion ratio reaching to 90.60% in sorghum. In total of 30.80 M reads and 133,246 SLAF labels were obtained and 319,428 SNPs were found. The associated region was located by SNP-index, Euclidean distance, and their combination. The candidate regions had a size of 1.95 Mb at nucleotides 54,788,026-56,740,873 on Chr.9. The SNPs locating at the associated region were analyzed between the two parents. Four non-synonymous-coding SNPs were found in this region. By verification, these SNPs were considered to be related to same height of tiller and main stem. Corresponding to three candidate genes (Sobic.009G197901.1, Sobic.009G213300.1, and Sobic.009G221200.1), these genes may be functional genes directly related to the traits.

Key words: sorghum, same height of tiller and main stem, SLAF, SNP

王瑞,凌亮,詹鹏杰,于纪珍,楚建强,平俊爱,张福耀. 控制高粱分蘖与主茎株高一致性的基因定位[J]. 作物学报, 2019, 45(6): 829-838.

Rui WANG,Liang LING,Peng-Jie ZHAN,Ji-Zhen YU,Jian-Qiang CHU,Jun-Ai PING,Fu-Yao ZHANG. Mapping of genes confessing same height of tiller and main stem in sorghum[J]. Acta Agronomica Sinica, 2019, 45(6): 829-838.

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染色体 Chromosome	染色体长度 Chromosome length	预测SLAF标签数 Expected SLAF number	平均SLAF间距 Average SLAF distance (bp)
Chr.1	84007898	11859	7083.89
Chr.2	88663305	12527	7077.78
Chr.3	84691115	11983	7067.61
Chr.4	77402132	10684	7244.68
Chr.5	70937750	9529	7444.41
Chr.6	70774438	9694	7300.88
Chr.7	73201405	10007	7315.02
Chr.8	63096686	8854	7126.35
Chr.9	67846938	9319	7280.50
Chr.10	69378333	9446	7344.73
合计Total	750000000	103902	7218.34

样品编号 Sample ID	总reads数 Total reads	Q30 (%)	GC (%)
父本 Male parent	4040595	92.05	45.96
母本 Female parent	3591789	92.49	45.96
混池aa Pool aa	10734806	92.02	45.58
混池ab Pool ab	12433150	90.24	45.67
水稻(对照) Rice (control)	192955	92.22	43.57

样品编号 Sample ID	SLAF标签数 SLAF number	测序总深度 Total depth	平均测序深度 Average depth
父本 Male parent	117244	2922769	24.93×
母本 Female parent	119317	2438178	20.43×
混池aa Pool aa	131993	8387924	63.55×
混池ab Pool ab	132818	8011244	60.32×

样品编号 Sample ID	SNP总数 Total SNP	SNP个数 SNP number	杂合率 Heterozygote ratio (%)
父本 Male parent	319428	221552	3.42
母本 Female parent	319428	233073	5.54
混池aa Pool aa	319428	265665	48.54
混池ab Pool ab	319428	312515	68.51

染色体 Chromosome	SLAF标签数 SLAF number	SNP数 SNP number
Chr.1	12565	31481
Chr.2	14026	40153
Chr.3	14064	35216
Chr.4	11159	24654
Chr.5	10947	29394
Chr.6	11411	31105
Chr.7	11939	24799
Chr.8	10144	25585
Chr.9	10634	27338
Chr.10	10933	25189
其他Other	15424	24514
合计Total	133246	319428

控制高粱分蘖与主茎株高一致性的基因定位

Mapping of genes confessing same height of tiller and main stem in sorghum

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染色体 Chromosome	关联区域起始位置 Start	关联区域终止位置 End	关联区域大小 Size (Mb)	关联区域内的基因数量 Gene number
Chr.9	30964929	38410608	7.45	44
Chr.9	43824552	59633348	15.81	1538
合计Total	—	—	23.26	1582

染色体 Chromosome	位置 Position	碱基替换 Transform	基因 Gene
Chr.9	55037659	C→A	Sobic.009G197901.1
Chr.9	55037906	T→G	Sobic.009G197901.1
Chr.9	56069487	T→G	Sobic.009G213300.1
Chr.9	56613839	A→G	Sobic.009G221200.1

引物名称 Primer ID	引物序列 Forward primer (5'-3')	引物序列 Reverse primer (5'-3')	产物大小 Size (bp)
SNP1	AAAATCACGTGGATAGGCCA	AGATCCGGCAATGATAGTGG	269
SNP2	CCACTATCATTGCCGGATCT	ATCTGAGGAAGCAAGCGAGA	233
SNP3	GGAGGAAATGCTGTGCTTCT	AAGCGATGGTCCATTGAATC	245
SNP4	TGATTCCTTCGTTGGAGGAG	TCACTAACATTGGGCAACCA	199

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