摘要
为了研究ADF基因对药用野生稻非生物胁迫的影响,利用药用野生稻干旱胁迫转录组数据分析的基础,通过PCR技术克隆OoADF1基因,序列分析结果表明,OoADF1的开放阅读框长度为453 bp,无内含子,编码150个氨基酸残基,具有典型的ADF结构域。qRT-PCR技术分析表明,OoADF1在四叶期的叶片、叶鞘和根,以及抽穗期的叶片、叶鞘、茎和幼穗等7个组织中均表达,四叶期根部表达量最高,抽穗期叶鞘表达量最高。干旱和高盐胁迫后,OoADF1表达量分别上调了17.9,40.0倍。将OoADF1完整的编码序列融合到原核表达载体p ET-28a(+)中,通过IPTG诱导及SDS-PASGE检测表明,OoADF1编码的蛋白分子量约为16 kDa,与预测的相符。OoADF1在大肠杆菌中的表达增加了大肠杆菌对高盐的抗性,表明OoADF1对高盐胁迫具有一定程度的响应能力,从而使得细胞免受高盐的毒害。
In order to investigate the effect of ADF gene on abiotic stress in Oryza officinalis.Based on the analysis on the drought-stressed transcriptome of Oryza officinalis,OoADF1 was cloned by RT-PCR technique and sequence analysis showed that OoADF1 contains 453 bp-length ORF and was intronless,and OoADF1 was predicted to contain 150 amino acids and have conserved ADF structure.Quantitative RT-PCR analysis showed that OoADF1 was expressed in all seven tissues,including leaf blade,leaf sheath and root at four-leaves stage,leaf blade,leaf sheath,stem and young panicle at heading stage in Oryza officinalis.The highest expression level of OoADF1 was detected in the root at four-leaves stage,while leaf sheath at heading stage.The fold of expression of OoADF1 amounted to 17.9 and 40.0 in the drought-or high Na Cl-stressed young seedling,respectively.The coding sequence of OoADF1 was fused to the prokaroytic expression vector p ET-28 a（＋）,and the combinant vectors were induced by IPTG and detected by SDS-PAGE.The molecular weight of protein OoADF1 was detected to be 16 kDa,which was consistent with theoretical one.The results of stress tolerance assay showed that the recombinant strain producing OoADF1 fusion protein improved the resistance to the high concentration,indicating that OoADF1 may exert a role in respond to salt stress and protect cells from high-salt harm.These results provides that OoADF1 has a certain degree of response to high salt stress,so that cells from high salt poisoning.
出处
《华北农学报》
CSCD
北大核心
2017年第6期37-44,共8页
Acta Agriculturae Boreali-Sinica
基金
国家自然科学基金项目(31271688)
