苦荞麦拒盐基因FtSOS1的克隆与表达研究

刘雪华*, 宋琎楠*, 陆启环*, 张玉喜, 侯丽霞, 于延冲, 张艳萍, 刘春英, 董春海, 杨洪兵**
青岛农业大学生命科学学院, 山东省高校植物生物技术重点实验室, 山东青岛266109

通信作者:刘雪华;E-mail: hbyang@qau.edu.cn

摘 要:

细胞质膜Na+/H+逆向转运蛋白基因SOS1在植物拒盐过程中发挥着十分重要的作用。本文以耐盐苦荞麦(Fagopyrum tataricum)品种‘川荞1号’及其新株系川荞1号-1为实验材料, 利用cDNA末端快速扩增(RACE)法获得苦荞麦SOS1基因, 命名为FtSOS1, 并在GenBank中注册, 登记号为KY659584。序列分析表明, 该基因全长3 873 bp, 开放阅读框3 396 bp, 编码1 131个氨基酸, 预测其蛋白分子质量为126 kDa, 等电点为6.57。系统进化树分析表明, FtSOS1与番茄(Lycopersicon esculentum) Le-SOS1蛋白、冰叶日中花(Mesembryanthemum crystallinum) McSOS1蛋白和胡杨(Populus euphratica) PeSOS1蛋白氨基酸同源率较高, 分别为61.35%、65.65%和64.17%。荧光定量PCR分析表明, 随着NaCl胁迫浓度的增加, FtSOS1基因在苦荞麦根部、茎基部和叶片的相对表达量显著增加, 150 mmol·L-1 NaCl胁迫下增加幅度最大, 分别比对照增加了304.10%、309.35%和260.08%。NaCl胁迫下, ‘川荞1号’和川荞1号-1茎基部FtSOS1基因均在6 h达到最大表达量, 叶片FtSOS1基因分别在6和12 h达到最大表达量, 说明苦荞麦FtSOS1基因的表达明显受盐胁迫诱导和调节。新株系川荞1号-1的FtSOS1基因表达量明显高于‘川荞1号’, 该基因的高表达可能是新株系耐盐性提高的重要原因, 这些结果与苦荞麦及其新株系耐盐生理指标的测定结果一致。

关键词:苦荞麦; 拒盐基因; 盐胁迫; FtSOS1基因克隆; FtSOS1基因表达

收稿:2017-06-13   修定:2017-10-26

资助:国家自然科学基金(31371552)。

Study on cloning and expression of salt exclusion gene FtSOS1 in tartary buckwheat

LIU Xue-Hua*, SONG Jin-Nan*, LU Qi-Huan*, ZHANG Yu-Xi, HOU Li-Xia, YU Yan-Chong, ZHANG Yan-Ping, LIU Chun-Ying, DONG Chun-Hai, YANG Hong-Bing**
Key Lab of Plant Biotechnology in Universities of Shandong / College of Life Sciences, Qingdao Agricultural University, Qingdao, Shandong 266109, China

Corresponding author: LIU Xue-Hua; E-mail: hbyang@qau.edu.cn

Abstract:

The cytoplasmic membrane Na+/H+ antiporter gene SOS1 plays an important role in the process of salt exclusion in plants. In this paper, the salt-tolerant tartary buckwheat (Fagopyrum tataricum) variety ‘Chuanqiao No. 1’ and its new line Chuanqiao No. 1-1 were used as experimental materials. The SOS1 gene of tartary buckwheat obtained by rapid amplification of cDNA ends (RACE) method was named FtSOS1, and was registered in GenBank under the accession number of KY659584. Sequence analysis showed that the total length of the gene is 3 873 bp, and the open reading frame is 3 396 bp, encoding 1 131 amino acids. The predicted protein molecular weight is 126 kDa and the isoelectric point is 6.57. Phylogenetic analysis showed that amino acids of FtSOS1 are high in homology rate with those of LeSOS1 (Lycopersicon esculentum), McSOS1 (Mesembryanthemum crystallinum) and PeSOS1 (Populus euphratica), which are 61.35%, 65.65% and 64.17%, respectively. Quantitative real-time PCR analysis showed that the relative expressions of FtSOS1 gene in roots, basal stem and leaf of tartary buckwheat were significantly increased with the concentration of NaCl stress increasing. The maxima were obtained under 150 mmol·L-1 NaCl stress, which were increased by 304.10%, 309.35% and 260.08%, respectively, in contrast with the control. Under NaCl stress, FtSOS1 gene of basal stem in ‘Chuanqiao No. 1’ and Chuanqiao No. 1-1 reached the maximal relative expressions after 6 h, and those of leaf in both plants reached the maxima after 6 and 12 h, respectively. It indicates that the expression of FtSOS1 gene of tartary buckwheat was obviously induced and regulated by salt stress, and the expression of FtSOS1 gene of new line Chuanqiao No.1-1 was obviously higher than that of ‘Chuanqiao No. 1’. The high expression of FtSOS1 gene might be an important cause to improve salt resistance of new line, and these results were consistent with physiological indexes of salt tolerance determined in tartary buckwheat and new line.

Key words: tartary buckwheat; salt exclusion gene; salt stress; FtSOS1 cloning; FtSOS1 expression

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