番茄苗期耐盐相关遗传位点鉴定及分子标记开发

doi:10.16420/j.issn.0513-353x.2023-0739

园艺学报 ›› 2024, Vol. 51 ›› Issue (2): 239-252.doi: 10.16420/j.issn.0513-353x.2023-0739

• 遗传育种·种质资源·分子生物学 • 上一篇下一篇

番茄苗期耐盐相关遗传位点鉴定及分子标记开发

徐琴¹, 王嘉颖¹, 张曼楠¹, 萧志浩¹, 郑涵楷¹, 卢永恩¹, 王涛涛¹, 张余洋¹^,², 张俊红¹^,², 叶志彪¹^,², 叶杰¹^,²^,^*()

¹ 华中农业大学园艺林学学院，果蔬园艺作物种质创新与利用全国重点实验室，武汉 430070
² 湖北洪山实验室，武汉 430070

收稿日期:2023-09-25 修回日期:2023-11-23 出版日期:2024-02-25 发布日期:2024-02-26
通讯作者:
*E-mail：yejie@mail.hzau.edu.cn
基金资助:
武汉市知识创新专项—曙光计划项目(2022020801020228); 中央高校基本科研业务费专项资金资助项目(2662022YJ014); 中央高校基本科研业务费专项资金资助项目(2662021YLQD003); 国家重点研发计划项目(2022YFF1003000); 华中农业大学高层次人才启动经费项目; 湖北洪山实验室重大项目(2021hszd007); 现代农业产业技术体系建设专项资助(CARS-23-A13)

Identification of Genetic Loci and Molecular Marker Development of Salt Tolerance in Tomato Seedlings

XU Qin¹, WANG Jiaying¹, ZHANG Mannan¹, XIAO Zhihao¹, ZHENG Hankai¹, LU Yong'en¹, WANG Taotao¹, ZHANG Yuyang¹^,², ZHANG Junhong¹^,², YE Zhibiao¹^,², YE Jie¹^,²^,^*()

¹ National Key Laboratory for Germplasm Innovation & Utilization of Horticultural Crops，College of Horticulture & Forestry Sciences，Huazhong Agricultural University，Wuhan 430070，China
² Hubei Hongshan Laboratory，Wuhan 430070，China

Received:2023-09-25 Revised:2023-11-23 Published:2024-02-25 Online:2024-02-26

摘要/Abstract

摘要：

以501份番茄重测序自然群体为研究材料，于苗期测定Na⁺和K⁺含量并计算Na⁺/K⁺比值，通过全基因组关联分析及单倍型分析，筛选耐盐相关显著关联位点，鉴定基于显著关联位点不同基因型材料的耐盐表型，利用该位点开发分子标记及验证。结果表明：大果番茄的驯化和改良过程导致了植株地上部Na⁺含量和Na⁺/K⁺比值的降低。通过全基因组关联分析在7号染色体上鉴定到与番茄地上部Na⁺含量和Na⁺/K⁺比值显著关联位点NTC7（Na⁺ tolerance loci on chromosome 7），该位点包含两个功能已知的耐盐基因，HKT1;1和HKT1;2。序列分析和单倍型分析鉴定到HKT1;2启动子上存在1个6 bp插入缺失与地上部中Na⁺含量和Na⁺/K⁺比值高度关联，通过盐处理验证该位点与番茄耐盐性相关，说明InDel_6是番茄苗期耐盐的功能突变位点。利用ARMS-PCR技术基于该位点开发番茄耐盐性相关的分子标记，并对不同基因型的材料进行耐盐表型鉴定，完成了分子标记的验证。

关键词: 番茄, 耐盐, 钠元素, 钾元素, 全基因组关联分析, 分子标记

Abstract:

To identify the genetic components of salt tolerance at the seedling stage of tomato，the contents of sodium（Na⁺）and potassium（K⁺）were determined，and genome-wide association analysis was further performed for shoot Na⁺/K⁺ ratio in a population consisting of 501 tomato accessions. The decrease in salt tolerance from wild tomatoes to large-fruited tomatoes is due to the decrease in Na⁺ content and Na⁺/K⁺ ratio during tomato domestication and improvement. The most significant loci associated with shoot Na⁺ content and Na⁺/K⁺ ratio were identified on chromosome 7，which contained two genes with known function，HKT1;1and HKT1;2. Sequence analysis，and haplotype analysis identified a 6 bp InDel（InDel_6）on the promoter of HKT1;2 showed highly correlated with Na⁺ content and Na⁺/K⁺ ratio，and salt treatment of different genotype accessions based on the InDel_6 confirmed that this mutation was the causative variant associated with Na⁺/K⁺ ratio and confer salt tolerance in tomato. Based on this causative variant，a codominant molecular marker that can quickly and accurately verify the salt tolerance of tomatoes at the seedling stage were developed.

Key words: tomato, salt tolerance, sodium, potassium, genome-wide association analysis, molecular marker

徐琴, 王嘉颖, 张曼楠, 萧志浩, 郑涵楷, 卢永恩, 王涛涛, 张余洋, 张俊红, 叶志彪, 叶杰. 番茄苗期耐盐相关遗传位点鉴定及分子标记开发[J]. 园艺学报, 2024, 51(2): 239-252.

XU Qin, WANG Jiaying, ZHANG Mannan, XIAO Zhihao, ZHENG Hankai, LU Yong'en, WANG Taotao, ZHANG Yuyang, ZHANG Junhong, YE Zhibiao, YE Jie. Identification of Genetic Loci and Molecular Marker Development of Salt Tolerance in Tomato Seedlings[J]. Acta Horticulturae Sinica, 2024, 51(2): 239-252.

图/表 6

图1 番茄3个亚组地上部和地下部Na+、K+含量及Na+/K+比值 PIM：醋栗番茄（46个）；CER：樱桃番茄（180个）；BIG：大果番茄（275个）。 * P < 0.05；** P < 0.001；*** P < 0.0001。

Fig. 1 The Na+ and K+ content and Na+/K+ ratio in roots and shoots of three subgroups of tomato accessions PIM：Solanum pimpinellifolium（46）；CER：S. lycopersicum var. cerasiforme（180）；BIG：S. lycopersicum（275）. * P < 0.05；** P < 0.01；*** P < 0.001.

图2 番茄种质资源地上部Na+含量、K+含量和Na+/K+比值全基因组关联分析结果

Fig. 2 Results of genome-wide association analysis of Na+ and K+ content in shoot of tomato germplasm resources

图3 Na+含量高度关联位点筛选及不同单倍型Na+含量、Na+/K+比值分析 A：钾离子转运家族成员SlHKT1;1和SlHKT1;2附近显著性位点分析；B：18 bp和6 bp插入缺失基因型Na+含量单倍型分析；C：Na+/K+比值单倍型分析，其中Hap_1：Del_18/Del_6，Hap_2：Ins_18/Del_6，Hap_3：Del_18/Ins_6，Hap_4：Ins_18/Ins_6；D、E：TS-9、TS-16、TS-50、TS-680、TS-17、TS-18、TS-123、TS-156材料InDel_18和InDel_6变异位点序列分析。

Fig. 3 Screening of highly associated loci of Na+ content and analysis of Na+ content and Na+/K+ ratio of different haplotypes A：Member of potassium ion transport family SlHKT1;1 and SlHKT1;2 analysis of nearby significant sites；B：Haplotype analysis of Na+ contentof 18 bp indel and 6 bp indel；C：Haplotype analysis of Na+/K+ ratio of four genotypes，Hap_1：Del_18/Del_6，Hap_2：Ins_18/Del_6，Hap_3：Del_18/Ins_6，Hap_4：Ins_18/Ins_6；D，E：Genotype sequencing results of 18 bp and 6 bp indel（TS-9，TS-16，TS-50，TS-680，TS-17，TS-18，TS-123，TS-156）.

图4 不同单倍型番茄盐处理的株高、根长和鲜质量相对变化及表型示例

Fig. 4 Relative changes in plant height，root length，and fresh mass and examples of phenotypes in tomato with different salt treated

图5 分子标记的电泳检测结果

Fig. 5 Examples of electrophoretic detection results of molecular markers

图6 不同单倍型番茄材料的分子标记检测结果

Fig. 6 Detection results of molecular markers in tomato materials with different haplotypes

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番茄苗期耐盐相关遗传位点鉴定及分子标记开发

Identification of Genetic Loci and Molecular Marker Development of Salt Tolerance in Tomato Seedlings

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番茄苗期耐盐相关遗传位点鉴定及分子标记开发

Identification of Genetic Loci and Molecular Marker Development of Salt Tolerance in Tomato Seedlings

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