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2025年4月3日 星期四
农业生物技术学报  2020, Vol. 28 Issue (7): 1230-1239    DOI: 10.3969/j.issn.1674-7968.2020.07.009
  研究论文与报告 本期目录 | 过刊浏览 | 高级检索 |
陆地棉胁迫相关蛋白基因GhSAP8的克隆及其耐盐性分析
王亦学1,2, 董艳辉1, 张欢欢1, 郝曜山1, 孙毅1, 吴慎杰1,2,*
1 山西省农业科学院 生物技术研究中心,太原 030031;
2 棉花种质资源利用与分子设计育种山西省重点实验室,运城 044000
Cloning and Salt-tolerance Analysis of Stress Associated Protein Gene GhSAP8 in Gossypium hirsutum
Wang Yi-Xue1,2, Dong Yan-Hui1, Zhang Huan-Huan1, Hao Yao-Shan1, Sun Yi1, Wu Shen-Jie1,2,*
1 Biotechnology Research Center, Shanxi Academy of Agricultural Sciences, Taiyuan 030031, China;
2 Shanxi Key Laboratory of Cotton Germplasm Resources Utilization and Molecular Design Breeding, Yuncheng 044000, China
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摘要 胁迫相关蛋白(stress associated proteins, SAPs)是具有A20结构域和(或) AN1结构域的一类锌指蛋白,广泛参与植物对逆境胁迫的响应。本研究通过同源克隆的方法获得陆地棉(Gossypium hirsutum) GhSAP8基因(GenBank No. XM_016813997.1),全长519 bp,编码172个氨基酸,预测蛋白分子量为18.22 kD,等电点为7.51。GhSAP8包含1个A20和1个AN1结构域,是典型的SAP蛋白结构域组合方式。qRT-PCR检测结果显示,GhSAP8基因在叶片中表达量最高,盐胁迫处理使该基因表达量增加,提示其可能参与盐胁迫响应。将GhSAP8基因构建到植物表达载体pCAMBIA1300并进行拟南芥(Arabidopsis thaliana)转化,在150 mmol/L NaCl处理条件下,过表达GhSAP8的拟南芥存活率(53%~70%)显著高于野生型对照(27%),表明过量表达该基因可提高拟南芥的耐盐能力。另外,在盐胁迫条件下,盐胁迫相关应答基因Δ1-吡咯啉-5-羧酸合成酶基因(Δ1-pyrroline-5-carboxylate synthetase 1, AtP5CS1)、脱水诱导基因(responsive to desiccation 29 B, AtRD29B)、盐超敏感基因(salt overly sensitive 1, AtSOS1)和Na+/H+转运蛋白基因 (sodium/hydrogen exchanger 1, AtNHX1)在转基因拟南芥中的基因表达量均显著高于野生型对照;而非盐胁迫条件下,转基因拟南芥中仅部分株系的AtP5CS1AtRD29BAtSOS1基因表达量高于野生型对照。上述结果提示,GhSAP8基因可能通过调控盐胁迫响应基因的表达来增强拟南芥的耐盐能力。本研究可为培育耐盐的转基因棉花提供候选基因。
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王亦学
董艳辉
张欢欢
郝曜山
孙毅
吴慎杰
关键词 棉花胁迫相关蛋白8基因(SAP8)基因克隆耐盐性    
Abstract:Stress associated proteins (SAPs) are zinc-finger proteins containing A20 and (or) AN1 domain which are involved in abiotic stress responses in plants. In this study, a stress associated protein gene, GhSAP8 (GenBank No. XM_016813997.1), was isolated from upland cotton (Gossypium hirsutum) via homology cloning. The full-length sequence of GhSAP8 gene was 519 bp, encoding a protein of 172 amino acids with a relative molecular weight of 18.22 kD and an isoelectric point of 7.51. GhSAP8 contained 1 conserved A20 domain in the N-terminus and 1 AN1 domain in the C-terminus which was typical SAP protein domain combinations. The expression of GhSAP8 gene was identified in different tissues, and qRT-PCR detection showed that the highest expression occurred in seedling leaf and it was induced by treatment of salt which suggested that GhSAP8 gene might be involved in responses to salt stress. The GhSAP8 gene was constructed into the plant expression vector pCAMBIA1300 and transformed into Arabidopsis thaliana. Under the treatment with 150 mmol/L NaCl, the survival rate (53%~70%) of Arabidopsis with overexpressed GhSAP8 gene was significantly higher than that of the wild type (27%), indicating the overexpression of GhSAP8 gene could enhance the tolerance to salt stress. In addition, the expression of salt-stress responsive genes, such as AtP5CS1 (Δ1-pyrroline-5-carboxylate synthetase 1), AtRD29B (responsive to desiccation 29 B), AtSOS1 (salt overly sensitive 1) and AtNHX1 (sodium/hydrogen exchanger 1), were significantly higher in transgenic Arabidopsis than that in wild type under salt stress. However, only a few of transgenic Arabidopsis lines showed increased expression of AtP5CS1, AtRD29B and AtSOS1 in wild type under non-salt stress. The above results indicated that GhSAP8 gene might improve tolerance to salt stress in transgenic Arabidopsis by regulating the expression of salt-stress responsive genes. The present study could provide superior gene resource for cultivation of transgenic salt-tolerant cotton.
Key wordsCotton    Stress associated protein 8 gene (SAP8)    Gene cloning    Salt tolerance
收稿日期: 2019-12-16     
ZTFLH:  S562  
基金资助:国家转基因生物新品种培育重大专项(2016ZX08005); 棉花种质资源利用与分子设计育种山西省重点实验室开放课题(MHKF201801)
通讯作者: * sj__wu@126.com   
引用本文:   
王亦学, 董艳辉, 张欢欢, 郝曜山, 孙毅, 吴慎杰. 陆地棉胁迫相关蛋白基因GhSAP8的克隆及其耐盐性分析[J]. 农业生物技术学报, 2020, 28(7): 1230-1239.
Wang Yi-Xue, Dong Yan-Hui, Zhang Huan-Huan, Hao Yao-Shan, Sun Yi, Wu Shen-Jie. Cloning and Salt-tolerance Analysis of Stress Associated Protein Gene GhSAP8 in Gossypium hirsutum. 农业生物技术学报, 2020, 28(7): 1230-1239.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/10.3969/j.issn.1674-7968.2020.07.009     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2020/V28/I7/1230
 
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