陆地棉纤维强度KASP-SNP标记的开发及效应评价

doi:10.13304/j.nykjdb.2022.0818

摘要/Abstract

摘要：

纤维强度（fiber strength，FS）是判断棉花纤维品质优劣的重要指标之一，开发可用于目标性状选择的分子标记将会大大提高选择准确性并加快育种进程。设计KASP引物，对国内外的376份品种（系）进行基因型鉴定，筛选多态性标记并验证其对高强纤维材料选择的有效性。3个SNP位点在材料中均具有多态性，标记FS-15对高强纤维材料的选择率为63.2%，FS-16和FS-29分别为60.0%和56.1%。对单倍型组合（Hap1、Hap2、Hap3和Hap4）的有效性评价表明，携带Hap3（TTA）的材料纤维强度显著高于其他组合，对高强纤维材料选择率达72.7%。此外，Hap3对纤维长度、铃重、籽指和衣指无不利影响。以上结果表明，开发的KASP标记及Hap3可用于高纤维强度材料的辅助选择，同时未对纤维长度、铃重、籽指和衣指等性状产生不利影响。

关键词: 棉花, 纤维强度, KASP-SNP标记, 单倍型组合, 效应评价

Abstract:

Fiber strength （FS） is an important indicator of fiber quality traits， and developing molecular markers associated with target traits would improve the selection accuracy and accelerate the process for breeding. The selected KASP markers were genotyped among 376 varieties （lines）， and then the polymorphism and the efficacy for higher strength were analyzed. 3 SNP markers showed polymorphic in the population， and the selection rate of FS-15 for high-strength fiber was 63.2% while FS-16 and FS-29 were 60.0% and 56.1%， respectively. Furthermore， the evaluation efficacy of different haplotype combination （Hap1， Hap2， Hap3 and Hap4） suggested that the accessions harboring Hap3 （TTA） showed higher fiber strength than those harboring other haplotypes. The selection rate of Hap3 for higher strength was 72.7%. In addition， Hap3 had no adverse effect on fiber length， boll weigh， seed index and lint index. The above results indicated these KASP markers and Hap3 could be used for selection of higher strength materials by providing genotypic information and did not adversely affect the fiber length， boll weight， seed index and lint index.

Key words: cotton, fiber strength, KASP-SNP markers, haplotype combination, effect evaluation

中图分类号:

S562

李丽花, 孙正文, 柯会锋, 谷淇深, 吴立强, 张艳, 张桂寅, 王省芬. 陆地棉纤维强度KASP-SNP标记的开发及效应评价[J]. 中国农业科技导报, 2024, 26(2): 46-55.

Lihua LI, Zhengwen SUN, Huifeng KE, Qishen GU, Liqiang WU, Yan ZHANG, Guiyin ZHANG, Xingfen WANG. Development and Effect Evaluation of KASP Markers for Fiber Strength in Gossypium hirsutum L.[J]. Journal of Agricultural Science and Technology, 2024, 26(2): 46-55.

图/表 12

表1 KASP分子标记引物序列

Table 1 Primer sequences used for KASP assays

标记 Marker	等位碱基 Allele	引物名称 Primer name	序列 Sequence （5’-3’）
FS-15	G/T	FS-15-FAM	TTGTTCTTCCAGACACCTTTGAAC
		FS-15-HEX	CTTTGTTCTTCCAGACACCTTTGAAA
		FS-15-Reverse	AGCTAAAAAATTGGAGGCTGAGAGGAAT
FS-16	A/T	FS-15-FAM	AAATTGAGCAAATTCAGGATGGAACCT
		FS-16-HEX	AAATTGAGCAAATTCAGGATGGAACCA
		FS-15-Reverse	ACATCAAAACAATCTGCTTGTACTGGCTT
FS-29	A/G	FS-29-FAM	ACGGAGCTGAAGGATGCAAGTAA
		FS-29-HEX	CGGAGCTGAAGGATGCAAGTAG
		FS-29-Reverse	GCAAACTCTCGATTTCAAATCCAAAGGTT

表2 376份材料在8个环境下纤维强度的变异分析

Table 2 Phenotypic variation for fiber strength of 376 accessions in 8 environments

年份Year	环境 Environment	最大值Max/（cN·tex^-1）	最小值Min/ （cN·tex^-1）	平均值Mean/ （cN·tex^-1）	标准差 SD	偏度 Skew	峰度 Kurt	变异系数 CV/%
2014	E1：保定Baoding	35.80	23.30	29.38	2.51	0.11	-0.045	8.54
	E2：河间Hejian	34.80	24.20	29.17	2.24	0.31	-0.26	7.68
	E3：辛集Xinji	36.30	25.20	30.80	2.21	0.068	-0.004	7.18
	E4：青县Qing county	36.80	24.80	30.08	2.16	0.16	0.066	7.18
	E5：崖城Yacheng	36.30	23.30	28.47	2.12	0.34	0.39	7.45
2015	E6：辛集Xinji	37.47	23.23	31.01	2.01	0.059	0.89	6.48
	E7：青县Qing county	35.51	19.91	28.39	2.34	0.044	0.29	8.24
	E8：崖城Yacheng	31.61	22.70	25.61	1.76	0.68	0.30	6.87

图1 376份陆地棉材料纤维强度频次分布

Fig. 1 FS frequency distribution of 376 upland cotton accessions

图2 引物FS-15在376份材料中的分型结果及对纤维强度的影响A：分型结果，每个圆点代表1个材料，红色、蓝色分别表示纯合基因型，绿色代表杂合型，粉色代表分型失败材料，黑点为空白对照；B：等位基因效应，*和**分别表示P<0.05和P<0.01水平差异显著，NA表示差异不显著；环境编号同表2

Fig. 2 Genotyping results of FS-15 in the population and allelic effects on fiber strengthA： Genotyping results of accessions， each dot represents 1 material，the red and blue represent homozygous， green represents heterozygous， pink represents failed data and black represents the non-template control， respectively； B： Allele effects， * and ** indicate significant difference at P<0.05 and P<0.01 levels， respectively， and NA indicates no significant difference； code of environment is same as Table 2

表3 KASP-SNP对纤维强度性状的影响

Table 3 Allelic effects of KASP-SNP markers on fiber strength

年份Year	环境 Environment	FS-15/ （cN·tex^-1）		效应值 Allele effect G<T	概率值 P-value	FS-16/ （cN·tex^-1）		效应值 Allele effect A<T	概率值 P-value	FS-29/ （cN·tex^-1）		效应值 Allele effect G<A	概率值 P-value
年份Year	环境 Environment	GG	TT	效应值 Allele effect G<T	概率值 P-value	AA	TT	效应值 Allele effect A<T	概率值 P-value	AA	GG	效应值 Allele effect G<A	概率值 P-value
2014	E1：保定Baoding	29.27	30.97	1.70	0.003 9	29.26	30.80	1.54	0.008 2	29.49	28.91	0.58	0.082 8
	E2：河间Hejian	29.06	30.32	1.26	0.017 8	29.12	30.21	1.08	0.036 0	29.27	28.83	0.44	0.139 7
	E3：辛集Xinji	30.67	32.27	1.61	0.002 0	30.74	32.11	1.36	0.006 7	30.81	30.63	0.18	0.543 5
	E4：青县Qing county	29.99	31.23	1.24	0.014 4	30.00	31.15	1.15	0.021 0	30.24	29.37	0.87	0.002 1
	E5：崖城Yacheng	28.29	29.98	1.69	0.000 5	28.38	29.84	1.46	0.002 9	28.58	27.93	0.65	0.020 9
2015	E6：辛集Xinji	30.98	31.14	0.15	0.753 9	30.97	31.09	0.12	0.798 9	31.03	30.92	0.11	0.693 7
	E7：青县Qing county	28.36	28.70	0.35	0.538 9	28.34	28.68	0.34	0.527 4	28.58	27.83	0.75	0.015 8
	E8：崖城Yacheng	25.54	26.44	0.90	0.031 4	25.56	26.39	0.83	0.041 7	25.76	24.98	0.78	0.000 7

图3 引物FS-16在376份材料中的分型结果及对纤维强度性状度的影响A：分型结果，每个圆点代表1个材料，红色、蓝色分别表示纯合基因型，绿色代表杂合型，粉色代表分型失败材料，黑点为空白对照；B：等位基因效应，*和**分别表示P<0.05和P<0.01水平差异显著，NA表示差异不显著；环境编号同表2

Fig. 3 Genotyping results of FS-16 in the population and allelic effects on fiber strengthA： Genotyping results of accessions， each dot represents 1 material，the red and blue represent homozygous， green represents heterozygous， pink represents failed data and black represents the non-template control， respectively； B： Allele effects， * and ** indicate significant difference at P<0.05 and P<0.01 levels， respectively， and NA indicates no significant difference； code of environment is same as Table 2

图4 引物FS-29在376份材料中的分型结果及对纤维强度性状度的影响A：分型结果，每个圆点代表1个材料，红色、蓝色分别表示纯合基因型，绿色代表杂合型，粉色代表分型失败材料，黑点为空白对照；B：等位基因效应，*和**分别表示P<0.05和P<0.01水平差异显著，NA表示差异不显著；环境编号同表2

Fig. 4 Genotyping results of FS-29 in the population and allelic effects on fiber strengthA： Genotyping results of accessions， each dot represents 1 material，the red and blue represent homozygous， green represents heterozygous， pink represents failed data and black represents the non-template control， respectively； B： Allele effects， * and ** indicate significant difference at P<0.05 and P<0.01 levels， respectively， and NA indicates no significant difference； code of environment is same as Table 2

表4 3个KASP-SNP标记对纤维强度性状的影响

Table 4 Allelic effects of 3 markers in the population of 376 accessions

基因

Gene

标记

Marker

等位基因（样本数）

Allele （number）

平均值Average/ （cN·tex^-1）

等位效应

Allele effect

选择率（样本数）

Selection rate/%（number）

概率值

P value

Gh_A07G1766（G/T）

FS-15

G（315）

T（19）

29.12

30.21

1.09

50.5（159）

63.2（12）

0.006 9

Gh_A07G1765 （A/T）

FS-16

A（336）

T（20）

29.15

30.10

0.95

51.8（174）

60.0（12）

0.016 0

Gh_D11G1931 （A/G）

FS-29

A（294）

G（70）

29.35

28.77

0.57

56.1（165）

40.0（28）

0.010 5

表5 单倍型组合

Table 5 Haplotype combination

标记Marker	Hap1	Hap2	Hap3	Hap4
FS-15（G/T）	G	G	T	T
FS-16（A/T）	A	A	T	T
FS-29（A/G）	A	G	A	G

图5 基于3个KASP标记构成的不同单倍型组合对强度性状的影响A：4种主要单倍型组合在376份材料出现频率；B：不同单倍型组合对纤维强度的影响，不同小写字母代表多重比较后显著性P<0.05

Fig. 5 Effects of different haplotype combination on fiber strength based on 3 KASP markersA： Frequency of four mainly haplotype combination in 376 accessions； B： Effect of different haplotype combianation on fiber strength， different lowercase letters represent Duncan’s multiple comparison at P<0.05 level

表6 不同单倍型组合对高强度纤维材料

Table 6 Effect of different haplotypes combination on accessions with high fiber strength

单倍型组合（样本数） Haplotype combination （number）	纤维强度 Fiber strength/（cN·tex^-1）	选择率（样本数） Selection rate /% （number）	概率值 P value
Hap1（242）	29.30±1.54 b	55.9（133）	1.62×10^-4
Hap2（54）	28.53±1.65 c	38.9（21）
Hap3（11）	30.56±1.73 a	72.7（8）
Hap4（7）	30.42±2.79 ab	57.1（4）

表 7 不同单倍型组合对纤维其他重要性状的影响

Table 7 Effects ofdifferent haplotype combinationon other traits

单倍型组合（样本数） Haplotype combination （number）	纤维长度 FL/mm	概率值 P value	铃重 BW/g	概率值 P-value	籽指 SI/g	概率值 P-value	衣指 LI/g	概率值 P value
Hap1（242）	29.18±0.99 b	5.45×10^-7	6.1±0.48 ab	0.028	11.1±0.83 ab	0.016	7.26±0.27 ab	0.021
Hap2（54）	28.37±1.14 c		5.93±0.43 b		10.91±0.90 b		7.08±0.67 b
Hap3（11）	29.71±0.90 a		6.29±0.42 a		11.59±1.04 a		7.74±1.01 a
Hap4（7）	29.3±1.30 ab		5.92±0.34 b		11.63±0.92 a		7.09±0.41 b

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