玉米ZmHsf04基因的克隆和特性及其对耐热性的调控

发布时间：2018-03-23 04:32

  本文选题：玉米　切入点：热激转录因子ZmHsf　出处：《农业生物技术学报》2017年09期 　论文类型：期刊论文

【摘要】：植物热激转录因子(heat shock transcription factor,Hsf)能够与热激蛋白基因启动子区域的热激元件结合而直接启动热激反应,因此成为热胁迫下基因转录激活信号转导通路中重要的调控因子。前人推测玉米(Zea mays)中至少有30个Hsf家族成员,其中A族18个,A2亚族4个。本实验室在前期对A1亚族Zm Hsf06结构、特性及其抗旱耐热性功能研究的基础上,采用同源克隆技术,从玉米幼叶中克隆获得A2亚族基因ZmHsf04(Gen Bank登录号:GRMZM2G010871_P01)的完整编码序列,序列全长1 074 bp,编码357个氨基酸残基。蛋白质结构包含Hsf家族DNA结合结构域,含有核定位信号(KRKELEDTISKKRRR)、核输出信号(LAQQLGYL)和激活结构域(LKMFESGVLN)等完整的功能结构域。蛋白序列与高粱(Sorghum bicolor)SORBIDRAFT_01g021490的同源性最高,达90%。荧光定量分析表明,正常生长条件下ZmHsf04在玉米多个组织器官中表达,幼嫩花粉中表达较高,约为幼嫩根系对照的16倍;ZmHsf04的表达水平受42℃热胁迫和外源脱落酸(abscisic acid,ABA)显著上调,最高表达量达正常对照的340倍,叶片峰值出现的时间分别为30 min和24 h;ZmHsf04也受水杨酸(SA)和H2O2上调表达,上调幅度低于热处理,最高表达量不超过12倍,显著低于热激或ABA处理,峰值出现相对滞后;用SA和H2O2分别预处理再热激,ZmHsf04的表达水平无明显差异。通过在洋葱(Allium cepa)表皮细胞中瞬时表达并观察绿色荧光蛋白(green fluorescence protein,GFP)荧光发现,ZmHsf04定位于细胞核。通过将ZmHsf04转化酵母(Saccharomyces cerevisiae)并进行50℃水浴热胁迫处理发现,ZmHsf04在酵母中可被半乳糖诱导蛋白表达,热胁迫同时降低正常和转ZmHsf04酵母的生长势,但与转空载体对照相比,转ZmHsf04基因酵母表现更强的耐热性。研究结果表明,ZmHsf04在植株花粉发育和热胁迫响应过程中可能发挥重要的调控功能。该研究为进一步解析ZmHsf04的功能及其调控机制提供理论和技术依据。
[Abstract]:The heat shock transcription factor Hsf) can bind to the heat shock element in the promoter region of the heat shock protein gene and initiate the heat shock reaction directly. Therefore, it has been suggested that there are at least 30 Hsf family members in Zea mays gene transduction pathway under heat stress, of which 4 belong to 18 A 2 subfamilies. On the basis of studying the function of drought resistance and heat resistance, the complete coding sequence of A2 subfamily gene ZmHsf04(Gen Bank was obtained from maize young leaves by homologous cloning technique, which was named as ZmHsf04(Gen Bank accession number: GRMZM2G010871P01). The sequence is 1 074 BP and encodes 357 amino acid residues. The protein structure contains the DNA binding domain of the Hsf family. Complete functional domains such as nuclear localization signal KRKELEDTISKRRRRN, nuclear output signal LQQLGYL) and activated domain LKMFESGVLN) were found. The homology of protein sequence with Sorghum bicolor)SORBIDRAFT_01g021490 was the highest, up to 90%. Under normal growth conditions, ZmHsf04 was expressed in several tissues and organs of maize, and the expression level of ZmHsf04 in young pollen was higher than that in young root control. The expression level of ZmHsf04 was significantly increased under 42 鈩，

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