Mining and Screening of Associated Markers and Candidate Genes Related to Seed Size and Shape in Cotton

doi:10.13304/j.nykjdb.2022.0341

Abstract

Abstract:

Cotton seed plays an important role in the processing of oil and fodder production， and also affects the cotton fiber yield and quality. In order to mine genetic loci and candidate genes related to the cotton seed， a natural population consisted of 419 cotton germplasms was assessed for the seed size and shape related traits in 2 years， and the genome wide association study （GWAS） was conducted by combining with the re-sequencing SNP genotypes. The results showed that there were relative wide genetic variations in the 5 seed related traits （seed length， seed width， seed area， seed perimeter and ratio of seed length to width） with the coefficient of variations 3.91%~9.55%. There were 7 SNPs associated with the seed size and shape in 2 years， and 23 SNPs associated with multiple related traits in single year. Among these SNPs， D11∶50561153 was associated with seed width， seed area and perimeter in two years， and D12∶56031535 was associated with seed length， seed area and perimeter in 2 years. Meanwhile， there were 4 SNPs on chromosome D05 associated with the ratio of seed length to width in 2 years. In addition， 10 SNPs on chromosome A07 were associated with seed length， seed area and seed perimeter， which including 2 haplotypes with significant differences on related traits. Further analysis showed that 21 candidate genes nearing the associated SNPs presented the increasing expression levels with the development of cotton ovule. Among these genes， the candidate gene Gh_D05G0148 encoded an EBF protein （EIN3-binding F box protein）， which was demonstrated to regulate the plant growth and development via the ethylene signal pathway. The expressions of Gh_D05G0148 showed much higher in the cotton ovule during 1~35 d post anthesis with the highest level at 10 d post anthesis. Meanwhile， the candidate gene Gh_D05G0144 was predicted to encode the ovule development involved YABBY transcription factor， and showed higher expression levels in cotton ovule during 20~35 d post anthesis. These results provided selection markers and candidate genes for the genetic improvements of the seed traits， and also laid foundation for the synchronous improvement of cotton fiber and seed traits.

Key words: cotton, seed size, seed shape, association analysis, candidate genes

摘要：

棉籽在油脂、饲料加工等领域具有重要用途，且影响纤维产量和品质。为发掘棉籽性状遗传位点与候选基因，利用419份具有丰富遗传变异的棉花品种资源构成的自然群体，在2个年度鉴定其棉籽性状，结合群体重测序SNP基因型进行全基因组关联分析。结果发现，供试群体棉籽长度、宽度、长宽比、面积与周长等性状存在较大遗传变异，变异系数3.91%~9.55%；有7个SNPs在2年同时被检测到与棉籽大小和形状关联，有23个SNPs在1年与3个以上棉籽相关性状关联，其中D11染色体D11∶50561153在2年与粒宽、面积和周长关联，D12染色体D12：56031535在2年与粒长、面积和周长关联；同时，D05染色体在2年检测到4个与籽粒长宽比显著关联SNPs，A07染色体检测到10个与粒长、周长和面积关联SNPs，且单倍型间相关性状差异极显著。进一步分析发现，关联位点附近存在21个候选基因随棉花胚珠发育进程表达量呈现增加趋势，其中Gh_D05G0148编码乙烯调控反应的EBF蛋白（EIN3-binding F-box protein），该基因在棉花开花后1~35 d的胚珠中优势表达，且以开花后10 d表达量最高；同时，候选基因Gh_D05G0144编码胚珠发育相关YABBY转录因子，该基因在棉花开花后20~35 d的胚珠中表达量较高，推测与棉籽后期发育有关。以上结果为棉籽多性状同步改良提供了选择标记与基因，为实现纤维和棉籽性状同步遗传改良奠定了基础。

关键词: 棉花, 棉籽大小, 棉籽形状, 关联分析, 候选基因

CLC Number:

S562

Huifeng KE, Zhengwen SUN, Guoning WANG, Chengsheng MENG, Liqiang WU. Mining and Screening of Associated Markers and Candidate Genes Related to Seed Size and Shape in Cotton[J]. Journal of Agricultural Science and Technology, 2022, 24(11): 76-86.

柯会锋, 孙正文, 王国宁, 孟成生, 吴立强. 棉籽大小与形状关联标记发掘及候选基因筛选[J]. 中国农业科技导报, 2022, 24(11): 76-86.

Figures/Tables 8

References 32

1	NIE X H, TU J L, WANG B, et al.. A BIL population derived from G . hirsutumG.barbadense provides a resource for cotton genetics and breeding [J/OL]. PLoS One, 2015, 10(10): e01410641 [2022-04-19]. .
2	LIU H Y, QUAMPAH A, CHEN J H, et al.. QTL mapping with different genetic systems for nine nonessential amino acids of cottonseeds [J]. Mol. Genet. Genomics, 2017, 292(10): 671-684.
3	SHANG L G, ABDUWELI A, WANG Y M, et al.. Genetic analysis and QTL mapping of oil content and seed index using two recombinant inbred lines and two backcross populations in upland cotton [J]. Plant Breeding, 2016, 135(2): 224-231.
4	钱玉源,刘祎,张曦,等.棉花种子质量突变体ims-15及其近等基因系种子发育期转录组分析[J].华北农学报, 2021, 36 (3): 50-59.
	QIAN Y Y, LIU Y, ZHANG X, et al.. Transcriptome analysis of cotton seed weight mutant ims-15 and its near isogenic line during seed development [J]. Acta Agric. Boreali-Sin., 2021, 36 (3): 50-59.
5	赵娟.调控内源生长素和细胞分裂素水平实现棉花种子纤维产量和品质的同步改良[D].重庆:西南大学, 2014.
	ZHAO J. Manipulation of auxin and cytokinin level in cotton concurrently increases fiber and seed yield [D]. Chongqing: Southwest University, 2014.
6	覃珊.胚中特异性下调细胞分裂素氧化酶基因(GhCKX)促进棉花种子发育[D].重庆:西南大学, 2009.
	QIN S. Ovule-specific down-regulation of GhCKX gene promoted seed development of cotton (Gossypium hirsutum L.) [D]. Chongqing: Southwest University, 2009.
7	王俊娟,叶武威,樊伟莉,等.不同基因型棉花种子活力的研究[J].种子科技, 2009, 27(9): 19-20.
	WANG J J, YE, W W, FAN W L, et al.. Study on vitality of different genotypes cotton seed [J]. Seed Sci. Technol., 2009, 27(9): 19-20.
8	ZHANG D, LI J P, COMPTON R O, et al.. Comparative genetics of seed size traits in divergent cereal lineages represented by sorghum (Panicoidae) and rice (Oryzoidae) [J]. G3: Genes Genom. Genet., 2015, 5(6): 1117-1128.
9	LI L, LI X H, LI L L, et al.. QTL identification and epistatic effect analysis of seed size and weight-related traits in Zea mays L [J/OL]. Mol. Breeding, 2019, 39(5): 67 [2022-04-20]. .
10	WILLIAMS K, SORRELLS M E. Three-dimensional seed size and shape QTL in hexaploid wheat (Triticum aestivum L.) populations [J]. Crop Sci., 2014, 54(1): 98-110.
11	CUI B F, CHEN L, YANG Y Q, et al.. Genetic analysis and map-based delimitation of a major locus qSS3 for seed size in soybean [J]. Plant Breeding, 2020, 139(2): 1145-1157.
12	HINA A, CAO Y C, SONG S Y, et al.. High-resolution mapping in two RIL populations refines major "QTL hotspot" regions for seed size and shape in soybean (Glycine max L.) [J/OL]. Int. J. Mol. Sci., 2020, 21(3): 1040 [2022-04-20]. .
13	GERAVANDI M, CHEGHAMIRZA K, FARSHADFAR E, et al.. QTL analysis of seed size and yield-related traits in an inter-gene pool population of common bean (Phaseolus vulgaris L.) [J/OL]. Scientia Hortic., 2020, 274(15): 109678 [2022-05-20]. .
14	SONG X J, HUANG W, SHI M, et al.. A QTL for rice grain width and weight encodes a previously unknown RING-type E3 ubiquitin ligase [J]. Nat. Genet., 2007, 39(5): 623-630.
15	程瑞如.小麦粒形的分子遗传分析及穗长基因HL1的精细定位[D].南京:南京农业大学, 2018.
	CHENG R R. Molecular genetic analysis of wheat kernel dimensions and fine mapping of spike length QTL HLl [D]. Nanjing: Nanjing Agricultural University, 2018.
16	耿庆河,王兰芬,武晶,等.普通菜豆籽粒大小与形状的QTL定位[J].作物学报,2017, 43(8): 1149-1160.
	GENG Q H, WANG L F, WU J, et al.. QTL mapping for seed size and shape in common bean [J]. Acta Agron. Sin., 2017, 43(8): 1149-1160.
17	秦伟伟,李永祥,李春辉,等.基于高密度遗传图谱的玉米籽粒性状QTL定位[J].作物学报, 2015, 41(10): 1510-1518.
	QIN W W, LI Y X, LI C H, et al.. QTL mapping for kernel related traits based on a high-density genetic map [J]. Acta Agron. Sin., 2015, 41(10): 1510-1518.
18	耿青青.大豆籽粒大小与形状性状的全基因组关联分析和DistortedMap软件包的研制[D].南京:南京农业大学, 2014.
	GENG Q Q. Genome-wide association studies of seed size and shape traits in soybean and development of distortedmap software [D]. Nanjing: Nanjing Agricultural University, 2014.
19	曾雅青.棉花种子活力的基因型差异及生态点影响[D].南京:南京农业大学, 2014.
	ZENG Y Q. Genotype difference and ecological environmental impact in cotton seed vigor [D]. Nanjing: Nanjing Agricultural University, 2014.
20	陈建华,杨继良,杨伟华,等.棉花种子活力及其影响因素探讨[J].棉花学报, 1995, 7(3): 134-140.
	CHEN J H, YANG J L, YANG W H, et al.. The study of cottonseed vigor and influence factors [J]. Acta Gossypii Sin., 1995, 7(3): 134-140.
21	MA Z Y, HE S P, WANG X F, et al.. Resequencing a core collection of upland cotton identifies genomic variation and loci influencing fiber quality and yield [J]. Nat. Genet., 2018, 50(6): 803-813.
22	GU Q S, KE H F, SUN Z W, et al.. Genetic variation associated with the shoot biomass of upland cotton seedlings under contrasting phosphate supplies [J/OL]. Mol. Breeding, 2020, 40(8): 80 [2022-04-20]. .
23	FANG Y X, ZHANG X Q, ZHANG X, et al.. A high-density genetic linkage map of SLAFs and QTL analysis of grain size and weight in barley (Hordeum vulgare L.) [J/OL]. Front. Plant Sci., 2020, 11: 620922 [2022-04-20]. .
24	LEI L, WANG L F, WANG S M, et al.. Marker-trait association analysis of seed traits in accessions of common bean (Phaseolus vulgaris L.) in China [J/OL]. Front. Genet., 2020, 11: 698 [2022-04-20]. .
25	ZHANG T Z, HU Y, JIANG W K, et al.. Sequencing of allotetraploid cotton (Gossypium hirsutum L. acc. TM-1) provides a resource for fiber improvement [J]. Nat. Biotechnol., 2015, 33(5): 531-537.
26	GUO X K, ZHANG Y, TU Y, et al.. Overexpression of an EIN3-binding F-box protein2-like gene caused elongated fruit shape and delayed fruit development and ripening in tomato [J]. Plant Sci., 2018, 272: 131-141.
27	陈秀玲,刘思源,孙婷,等.番茄YABBY基因家族的生物信息学分析[J].东北农业大学学报, 2017, 48(10): 11-19.
	CHEN X L, LIU S Y, SUN T, et al.. Bioinformatics analysis on YABBY gene family in tomato [J]. J. Northeast Agric. Univ., 2017, 48(10): 11-19.
28	ZHANG M, WANG C P, LIN Q F, et al.. A tetratricopeptide repeat domain-containing protein SSR1 located in mitochondria is involved in root development and auxin polar transport in Arabidopsis [J]. Plant J., 2015, 83(4): 582-599.
29	WEI K F, LI Y X. Functional genomics of the protein kinase superfamily from wheat [J/OL]. Mol. Breeding, 2019, 39: 141 [2022-04-20]. .
30	WANG W W, SUN Y, YANG P, et al.. A high density SLAF-seq SNP genetic map and QTL for seed size, oil and protein content in upland cotton [J/OL]. BMC Genomics, 2019, 20(1): 599 [2022-04-20]. .
31	FINET C, FLOYD S K, CONWAY S J, et al.. Evolution of the YABBY gene family in seed plants [J]. Evol. Dev., 2016, 18(2): 116-126.
32	SUN L, WEI Y Q, WU K H, et al.. Restriction of iron loading into developing seeds by a YABBY transcription factor safeguards successful reproduction in Arabidopsis [J]. Mol. Plant, 2021, 14(10): 1624-1639.

性状 Trait	年度 Year	均值 Mean	标准差 SD	变异系数 CV/%	最大值 Max.	最小值 Min.	偏度系数 Skew	峰度系数 Kurt
粒长 Seed length/mm	2015	8.71	0.49	5.63	10.34	7.44	0.48	0.49
粒长 Seed length/mm	2017	8.20	0.45	5.49	10.54	7.09	0.59	1.52
粒宽 Seed width/mm	2015	4.74	0.23	4.85	5.46	4.06	0.16	0.30
粒宽 Seed width/mm	2017	4.59	0.22	4.79	5.67	3.92	0.55	1.64
籽粒长宽比 Ratio of seed length to width	2015	1.85	0.08	4.32	2.11	1.65	0.41	0.50
籽粒长宽比 Ratio of seed length to width	2017	1.79	0.07	3.91	2.04	1.57	0.26	1.04
籽粒面积 Seed area/mm²	2015	30.89	2.92	9.45	42.11	23.76	0.48	0.57
籽粒面积 Seed area/mm²	2017	28.69	2.74	9.55	45.16	22.38	0.89	3.07
籽粒周长 Seed perimeter/mm	2015	22.81	1.21	5.30	28.27	20.11	0.59	1.09
籽粒周长 Seed perimeter/mm	2017	21.43	1.10	5.13	27.32	18.96	0.65	1.71

性状 Trait	年度 Year	均值 Mean	标准差 SD	变异系数 CV/%	最大值 Max.	最小值 Min.	偏度系数 Skew	峰度系数 Kurt
粒长 Seed length/mm	2015	8.71	0.49	5.63	10.34	7.44	0.48	0.49
粒长 Seed length/mm	2017	8.20	0.45	5.49	10.54	7.09	0.59	1.52
粒宽 Seed width/mm	2015	4.74	0.23	4.85	5.46	4.06	0.16	0.30
粒宽 Seed width/mm	2017	4.59	0.22	4.79	5.67	3.92	0.55	1.64
籽粒长宽比 Ratio of seed length to width	2015	1.85	0.08	4.32	2.11	1.65	0.41	0.50
籽粒长宽比 Ratio of seed length to width	2017	1.79	0.07	3.91	2.04	1.57	0.26	1.04
籽粒面积 Seed area/mm²	2015	30.89	2.92	9.45	42.11	23.76	0.48	0.57
籽粒面积 Seed area/mm²	2017	28.69	2.74	9.55	45.16	22.38	0.89	3.07
籽粒周长 Seed perimeter/mm	2015	22.81	1.21	5.30	28.27	20.11	0.59	1.09
籽粒周长 Seed perimeter/mm	2017	21.43	1.10	5.13	27.32	18.96	0.65	1.71

序号 No.	染色体 Chromosome	SNP	物理位置 Physical position	等位变异Allele	性状 Trait	年度 Year	-log₁₀P
5	D05	D05∶1496927	1 496 927	T/C	RSLW	2015， 2017	5.21， 5.34
6	D05	D05∶1512276	1 512 276	T/A	RSLW	2015， 2017	5.92， 5.25
7	A04	A04∶54398482	54 398 482	C/A	SP	2015， 2017	5.09， 5.32
					SA	2017	5.22
8#	A07	A07∶70678488	70 678 488	G/C	SL， SA，SP	2015	6.89， 6.69， 6.72
9#	A07	A07∶70678557	70 678 557	A/G	SL，SA，SP	2015	5.72， 5.35， 5.40
10#	A07	A07∶70679600	70 679 600	T/C	SL，SA，SP	2015	5.18， 5.49， 5.27
11#	A07	A07∶70679917	70 679 917	C/T	SL，SA，SP	2015	6.27， 5.87， 5.96
12#	A07	A07∶70680268	70 680 268	A/C	SL，SA，SP	2015	6.14， 5.83， 5.84
13#	A07	A07∶70680314	70 680 314	A/G	SL，SA，SP	2015	6.45， 6.38， 6.39
14#	A07	A07∶70682150	70 682 150	C/A	SL，SA，SP	2015	5.87， 5.22， 5.51
15#	A07	A07∶70682969	70 682 969	T/C	SL，SA，SP	2015	5.25， 5.19， 5.29
16#	A07	A07∶70700642	70 700 642	C/T	SL，SA，SP	2015	5.56， 5.02， 5.16
17	A07	A07∶70706541	70 706 541	C/G	SL，SA，SP	2015	6.03， 5.21， 5.66
18	A07	A07∶72096450	72 096 450	A/G	RSLW，SL，SP	2015	5.10， 6.10， 5.12
19	A07	A07∶44855893	44 855 893	G/A	SL，SA，SP	2015	5.98， 6.06， 6.22
20	A08	A08∶28488249	28 488 249	T/C	SL，SA，SP	2015	5.17， 5.68， 5.64
21	A08	A08∶32230490	32 230 490	T/C	SL，SA，SP	2015	5.41， 5.48， 5.28
22	A08	A08∶97095613	97 095 613	G/C	SL，SA，SP	2015	5.23， 5.45， 5.76
23	A01	A01∶1540366	1 540 366	C/T	SL，SA，SP	2017	5.80， 5.25， 5.09
24	A01	A01∶12448791	12 448 791	A/G	SL，SW，SA，SP	2015	5.00， 5.89， 6.43， 5.85
25	A02	A02∶6005362	6 005 362	C/T	SL，SA，SP	2015	6.51， 5.03， 6.07
26	A02	A02∶10265596	10 265 596	G/A	SL，SA，SP	2017	6.20， 5.36， 5.90
27	A05	A05∶20705321	20 705 321	C/G	SW，SA，SP	2017	5.06， 5.04， 5.39
28	A05	A05∶37081549	37 081 549	A/G	SL，SA，SP	2017	6.63， 5.06， 6.17
29	A06	A06∶70904173	70 904 173	A/G	SL，SA，SP	2015	5.07， 5.05， 5.11
30	D08	D08∶34053083	34 053 083	T/C	SL，SA，SP	2017	5.90， 5.73， 5.74

序号 No.	染色体 Chromosome	SNP	物理位置 Physical position	等位变异Allele	性状 Trait	年度 Year	-log₁₀P
5	D05	D05∶1496927	1 496 927	T/C	RSLW	2015， 2017	5.21， 5.34
6	D05	D05∶1512276	1 512 276	T/A	RSLW	2015， 2017	5.92， 5.25
7	A04	A04∶54398482	54 398 482	C/A	SP	2015， 2017	5.09， 5.32
					SA	2017	5.22
8#	A07	A07∶70678488	70 678 488	G/C	SL， SA，SP	2015	6.89， 6.69， 6.72
9#	A07	A07∶70678557	70 678 557	A/G	SL，SA，SP	2015	5.72， 5.35， 5.40
10#	A07	A07∶70679600	70 679 600	T/C	SL，SA，SP	2015	5.18， 5.49， 5.27
11#	A07	A07∶70679917	70 679 917	C/T	SL，SA，SP	2015	6.27， 5.87， 5.96
12#	A07	A07∶70680268	70 680 268	A/C	SL，SA，SP	2015	6.14， 5.83， 5.84
13#	A07	A07∶70680314	70 680 314	A/G	SL，SA，SP	2015	6.45， 6.38， 6.39
14#	A07	A07∶70682150	70 682 150	C/A	SL，SA，SP	2015	5.87， 5.22， 5.51
15#	A07	A07∶70682969	70 682 969	T/C	SL，SA，SP	2015	5.25， 5.19， 5.29
16#	A07	A07∶70700642	70 700 642	C/T	SL，SA，SP	2015	5.56， 5.02， 5.16
17	A07	A07∶70706541	70 706 541	C/G	SL，SA，SP	2015	6.03， 5.21， 5.66
18	A07	A07∶72096450	72 096 450	A/G	RSLW，SL，SP	2015	5.10， 6.10， 5.12
19	A07	A07∶44855893	44 855 893	G/A	SL，SA，SP	2015	5.98， 6.06， 6.22
20	A08	A08∶28488249	28 488 249	T/C	SL，SA，SP	2015	5.17， 5.68， 5.64
21	A08	A08∶32230490	32 230 490	T/C	SL，SA，SP	2015	5.41， 5.48， 5.28
22	A08	A08∶97095613	97 095 613	G/C	SL，SA，SP	2015	5.23， 5.45， 5.76
23	A01	A01∶1540366	1 540 366	C/T	SL，SA，SP	2017	5.80， 5.25， 5.09
24	A01	A01∶12448791	12 448 791	A/G	SL，SW，SA，SP	2015	5.00， 5.89， 6.43， 5.85
25	A02	A02∶6005362	6 005 362	C/T	SL，SA，SP	2015	6.51， 5.03， 6.07
26	A02	A02∶10265596	10 265 596	G/A	SL，SA，SP	2017	6.20， 5.36， 5.90
27	A05	A05∶20705321	20 705 321	C/G	SW，SA，SP	2017	5.06， 5.04， 5.39
28	A05	A05∶37081549	37 081 549	A/G	SL，SA，SP	2017	6.63， 5.06， 6.17
29	A06	A06∶70904173	70 904 173	A/G	SL，SA，SP	2015	5.07， 5.05， 5.11
30	D08	D08∶34053083	34 053 083	T/C	SL，SA，SP	2017	5.90， 5.73， 5.74

序号 No.	染色体 Chromosome	SNP	物理位置 Physical position	等位变异Allele	性状 Trait	年度 Year	-log₁₀P
1	D11	D11∶50561153	50 561 153	C/T	SW	2015， 2017	5.32， 6.53
					SA	2015， 2017	6.06， 6.45
					SP	2015， 2017	5.81， 5.76
2	D12	D12∶56031535	56 031 535	A/C	SL	2015， 2017	5.85， 5.35
					SP	2015， 2017	5.33， 5.46
					SA	2015	5.16
3	D05	D05∶1490099	1 490 099	G/A	RSLW	2015， 2017	8.00， 6.83
4	D05	D05∶1490516	1 490 516	G/T	RSLW	2015， 2017	5.63， 5.36