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利用RNA干涉技术研究水稻锌指蛋白基因O_sBBX22的生物学功能,为探讨O_sBBX22响应热胁迫的机制、培育抗逆水稻、减轻高温对水稻的损害奠定基础。通过观察转基因突变体植株和野生型植株在热胁迫下的表型差异,分析O_sBBX22生物学功能;采用半定量PCR和荧光定量PCR检测O_sBBX22以及相关的热激转录因子(HSF)、热激蛋白(HSP)基因在转基因突变体株系中的表达水平;通过原位组织化学检测过氧化氢在野生型、转基因突变体株系叶片中的定位和积累情况。结果表明,在05 h热胁迫条件下,与野生型株系相比,O_sBBX22的表达在转基因突变体植株中明显下调;而野生型O_sBBX22受热信号诱导,随着热激时间的增加,O_sBBX22的表达量呈先上升后下降的趋势,且在热激1 h时表达量最高。相关的HSF和HSP也受热信号诱导,野生型株系中的HSFA2a、HSFA7、HSP16.9和HSP100表达量均比转基因突变体株系高,且在热激1 h时,HSFA2a、HSP16.9和HSP100表达量最高,而HSFA7在热激3 h时表达最高。热胁迫3 h,经DAB染色,转基因突变体株系叶片上出现的红褐色斑点主要集中于叶脉和受损伤部位,且明显多于野生型。锌指蛋白基因O_sBBX22在水稻苗期热胁迫应答中具有重要的作用,野生型株系抗热能力明显高于O_sBBX22抑制表达转基因株系;HSFA2a、HSFA7、HSP16.9和HSP100可能参与了O_sBBX22介导的水稻耐热调控。
Abstract：Biological function of rice zinc-finger protein gene O_sBBX22 was investigated by RNA interference,which laid a foundation for exploring the mechanism of O_sBBX22 response to heat stress, developing stress tolerance rice, and reducing the damage to rice caused by heat stress. The biological functions of O_sBBX22 were analyzed by observing the phenotypic differences between transgenic mutant plants and wild type plants underheatstress. Semi-quantitative PCR and fluorescence quantitative PCR were used to detect the expression level of O_sBBX22 and its related heat shock transcription factor(HSF), heat shock protein(HSP) gene in transgenic mutant lines. In situ histochemistry was used to detect the location and accumulation of hydrogen peroxide in leaves of wild-type and transgenic mutants. The results showed that the expression level of O_sBBX22 in transgenic line was distinctly down-regulated compared with wild lines under the condition of 05 h heat stress, while O_sBBX22 in wild plants could be induced by heat signals and the expression level of O_sBBX22 increased first and then decreased with the rise of heat shock time, and it reached the highest after 1 hour under heat stress. The related HSF and HSP were also induced by heat signals and the expression levels of HSFA2 a, HSFA7, HSP16.9 and HSP100 in wild plants were higher than those in transgenic plants. The expression levels of HSFA2 a, HSP16.9 and HSP100 were the highest after 1 hour under heat stress, but that of HSFA7 reached peak after 3 hours under heat stress. After 3 hours under heatstress, the reddish brown spots on the leaves of transgenic plants were mainly located in veins and damaged parts which were significantly more than those in thewild type by DAB staining.Zinc finger protein gene O_sBBX22 might play an important role in response to heat stress in rice seedlings, and the heat resistance of wild type lines was significantly higher than that of O_sBBX22 inhibited expression transgenic lines. HSFA2 a, HSFA7, HSP16.9 and HSP100 might be involved in the regulation of heat tolerance in rice.
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基本信息:
DOI：10.13417/j.gab.037.000836
中图分类号:Q943.2;S511
引用信息:
[1]骆鹰,谢旻,张超,等.水稻锌指蛋白基因O_sBBX22响应热胁迫的功能分析[J].基因组学与应用生物学,2018,37(02):836-844.DOI:10.13417/j.gab.037.000836.
基金信息:
国家自然基金(31301081);; 科技部国家科技支撑计划项目(2014BAD01B04);; 教育部博士点基金项目(2013-4320120009);; 湖南省高校创新平台开放基金项目(13K063);; 湘南优势植物资源综合利用湖南省重点实验室资助项目(湘南植物[2015]1号)共同资助
2017-06-12
2017-06-12
2017-06-12