大豆GmGBP1基因对转基因烟草幼苗耐热性的影响

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摘要大豆中受赤霉素诱导的MYB类转录因子结合蛋白1(Glycine max gibberellin-regulated Myb-related transcription factor binding protein 1, GmGBP1)基因是斯隆-凯特琳逆转录病毒互作蛋白(Sloan-Kettering retrovirus interacting protein, SKIP)基因的同源基因。GmGBP1编码的蛋白质不仅参与调控多种植物的非生物胁迫信号转导途径，同时还参与脱落酸、赤霉素等植物激素的信号转导。本研究将大豆GmGBP1基因转化烟草(Nicotiana tabacum)，并使该基因在烟草中过量表达。结果表明，经过48 ℃处理48 h后，野生型烟草(wild-type tobacco, WT)幼苗叶片萎蔫坏死、失绿变黄；过表达大豆GmGBP1基因的转基因烟草(transgenic tobacco of overexpression GmGBP1 gene, GBP1ox)幼苗叶片未出现明显变黄。55 ℃高温处理12 h后，室温恢复生长15 d，GBP1ox烟草幼苗存活率为24.14%，高于WT。经48 ℃处理48 h后，WT烟草幼苗叶片中的丙二醛(malondialdehyde, MDA)含量是GBP1ox的2.07倍；用二氨基联苯胺(diaminobenzidine, DAB)对高温处理后的烟草幼苗叶片染色，WT烟草幼苗叶片染色面积占叶片总面积的比例为43.66%，显著高于GBP1ox(P＜0.05)。对烟草幼苗55 ℃高温处理3 、6 、7和8 h时，GBP1ox烟草幼苗叶片中拟南芥锌指结构蛋白12 (zinc finger protein of Arabidopsis thaliana 12, ZAT12)基因的相对表达量显著高于WT(P＜0.05)。但是高温处理后WT与GBP1ox烟草幼苗叶片中热休克蛋白40 (heat shock protein 40, HSP40)基因的相对表达量没有明显差异。上述结果表明，过量表达大豆GmGBP1基因能在一定程度上提高转基因烟草幼苗的耐热性。本研究为耐热转基因作物新品种的培育提供参考，并为进一步研究大豆GmGBP1基因的功能提供依据。

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	聂腾坤
	赵琳
	李文滨
	丁福全
	李敏敏
	杨雪
	张可欣
	孙晶哲
	杜海龙

关键词 ：大豆, 烟草幼苗, 大豆中受赤霉素诱导的MYB类转录因子结合蛋白基因(GmGBP1), 斯隆-凯特琳逆转录病毒互作蛋白基因(SKIP), 耐热

Abstract：Soybean (Glycine max) gibberellin-regulated MYB-related transcription factor binding protein 1 (GmGBP1) gene is a homologous gene of Sloan-Kettering retrovirus interacting protein (SKIP) gene . Proteins encoded by GmGBP1 not only participate in the regulation of a variety of abiotic stress signal transduction pathways in plants, but also play a role in the signal transduction of plant hormones, such as abscisic acid and gibberellin. In this study, soybean GmGBP1 gene was transformed into transgenic tobacco (Nicotiana tabacum) and overexpressed. After being treated at a high temperature of 48℃ for 48 h, leaf wilting and necrosis, leaf chlorosis and leaf etiolation were detected in the wild-type (WT) tobacco seedlings; while no obvious yellowing was found in the leaves of transgenic tobacco of overexpression GmGBP1 gene (GBP1ox) seedlings at a high temperature. After being treated at a high temperature of 55 ℃ for 12 h, tobacco seedlings were placed at room temperature for recovery growth for 15 d, and the survival rate of over-expression of GmGBP1 gene in tobacco seedlings was 24.14%. The survival rate of GBP1ox tobacco seedling was higher than the WT tobacco seedlings. The malondialdehyde (MDA) content of WT leaf tobacco seedlings was 2.07-fold more than that in GBP1ox after 48 ℃ treatment for 48 h. The leaves of tobacco seedlings were stained by using diaminobenzidine (DAB) after being treated at a high temperature, and the staining results demonstrated that the ratio of stained area of WT tobacco seedling leaves to total leaf area was 43.66%. The percentage of the total leaf area of DAB staining in WT seedling tobacco was significantly greater than GBP1ox tobacco seedlings (P＜0.05) as a result. High temperature treatment was performed in tobacco seedlings to analyze the changes in the relative expression levels of heat-resistance related genes in tobaccos. Zinc finger protein of Arabidopsis thaliana 12 (ZAT12) gene expression was significantly higher in GBP1ox leaf tobacco seedlings than that in WT (P＜0.05) for 3, 6, 7 and 8 h treatment at 55 ℃ high temperature. On the other hand, there was no significant difference in the relative expression levels of heat shock protein 40 (HSP40) gene in WT and GBP1ox the leaves of tobacco seedlings after being treated at a high temperature. The results showed that overexpression of GmGBP1 gene could improve the heat resistance of transgenic tobacco seedlings to some extent. The present study provides references for the cultivation of new heat-resistant transgenic crops, and lays a solid foundation on further investigations about the functions of soybean GmGBP1 gene.

Key words： Soybean Tobacco seedling Glycine max gibberellin-regulated Myb-related transcription factor binding protein gene(GmGBP1), Sloan-Kettering retrovirus interacting protein gene(SKIP) Heat tolerance

收稿日期: 2015-10-08 出版日期: 2016-02-05

基金资助:国家自然科学基金;转基因生物新品种培育科技重大专项;东北农业大学“青年才俊”项目;黑龙江省自然科学基金;黑龙江省教育厅青年学术骨干项目;十二五农村领域国家科技计划课题;东北农业大学研究生科技创新基金资助

通讯作者: 李文滨 E-mail: wenbinli@yahoo.com

引用本文:

聂腾坤赵琳李文滨丁福全李敏敏杨雪张可欣孙晶哲杜海龙. 大豆GmGBP1基因对转基因烟草幼苗耐热性的影响[J]. , 2016, 24(3): 313-322.

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http://journal05.magtech.org.cn/Jwk_ny/CN/ 或 http://journal05.magtech.org.cn/Jwk_ny/CN/Y2016/V24/I3/313

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