割手密抗旱相关基因SsREMO-1a的表达与功能分析  

Expression and functional analysis of drought resistance related gene SsREMO-1a in Saccharum spontaneum L.

作　　者：洪亚楠 刘洋 姚艳丽[2] 胡小文 徐磊[3] HONG Ya-nan;LIU Yang;YAO Yan-li;HU Xiao-wen;XU Lei(Jiaxing Vocational and Technical College,Jiaxing,Zhejiang 314036,China;Institute of South Subtropical Crops,Chinese Academy of Tropical Agricultural Sciences,Zhanjiang,Guangdong 524013,China;Zhanjiang Experimental Station,Chinese Academy of Tropical Agricultural Sciences,Zhanjiang,Guangdong 524013,China)

机构地区：[1]嘉兴职业技术学院,浙江嘉兴314036 [2]中国热带农业科学院南亚热带作物研究所,广东湛江524013 [3]中国热带农业科学院湛江实验站,广东湛江524013

出　　处：《西南农业学报》2024年第6期1196-1202,共7页Southwest China Journal of Agricultural Sciences

基　　金：嘉兴职业技术学院“三创”项目(SCXM202201);嘉兴职业技术学院校级科研项目(jzyz202206);国家重点研发计划项目子课题(2018YFD1000503);中国职业技术教育学会科研课题(ZJ2023B117)。

摘　　要：【目的】获得割手密SsREMO-1a基因全序列,开展组织特异表达、qPCR分析、亚细胞定位研究,研究转SsREMO-1a基因拟南芥的表型变化以及CAT基因的表达模式。【方法】以海南甘蔗野生种(割手密Sp-24)为材料,通过PCR、纯化、克隆和测序验证后获得完整的割手密SsREMO-1a基因序列。对苗期正常生长的割手密进行干旱胁迫处理,于胁迫处理后0、1、2、3、4和5 d收集幼嫩叶片、根和茎等组织进行组织特异表达分析和qPCR分析。采用烟草亚细胞定位技术、基因遗传转化技术等对SsREMO-1a的表达情况进行组织定位和功能分析。构建pBI221-SsREMO-1a-GFP融合表达载体,通过农杆菌转化烟草进行细胞定位分析。构建过量表达载体pCAMBIA 1304-SsREMO-1a,利用农杆菌转化法将SsREMO-1a导入拟南芥,观测干旱胁迫后0、1、2、3、4和5 d植株表型以及CAT-1基因表达量。【结果】SsREMO-1a基因全长均为738 bp,开放读码框(ORF)564 bp,编码187个氨基酸;组织特异性表达表明,SsREMO-1a在根、茎、叶中均有表达;qPCR分析结果表明,该基因受干旱胁迫的诱导并呈显著上调表达;亚细胞定位结果表明,SsREMO-1a蛋白定位于细胞膜上;共获得11个转基因拟南芥株系,抗旱性鉴定表明转基因植株能够提高植株的抗旱能力;转基因植株CAT-1基因表达量显著升高。【结论】SsREMO-1a能够提高植株的抗旱能力,对CAT-1基因具有一定的调控作用。【Objective】The present paper aimed to obtain the complete sequence of SsREMO-1a gene,conduct tissue-specific expression,qPCR expression analysis,sub-cellular localization study,and the phenotypic changes of Arabidopsis thaliana with SsREMO-1a gene transfer and the expression pattern of CAT-1 gene.【Method】The complete sequence of SsREMO-1a gene was obtained by PCR,purification,cloning and sequencing from Hainan sugarcane wild species(Sp-24).Drought stress was applied to the normal growth of the seedlings.Young leaves,roots and stems at 0,1,2,3,4 and 5 days after the stress treatment were collected for tissue-specific expression analysis and qPCR analysis.The tissue location and function of SsREMO-1a were analyzed by tobacco sub-cellular mapping and gene transformation techniques.pBI221-SsREMO-1a-GFP fusion expression vector was constructed and transformed into tobacco for cell localization analysis.The over-expression vector pCAMBIA 1304-SsREMO-1a was constructed,and SsREMO-1a was introduced into A.thaliana by agrobacterium transformation.The phenotypes of plants at 0,1,2,3,4 and 5 days after drought stress and the expression of CAT-1 gene were observed.【Result】The full length of SsREMO-1a gene was 738 bp,and the open code frame(ORF)was 564 bp,encoding 187 amino acids.Tissue-specific expression showed that SsREMO-1a was expressed in roots,stems and leaves.qPCR results showed that the gene was induced by drought stress and showed significantly up-regulated expression.The results of sub-cellular localization showed that SsREMO-1a protein was localized on the cell membrane.A total of 11 transgenic A.thaliana lines were obtained,and the drought resistance identification showed that the transgenic plants could improve the drought resistance of the plants.The expression of CAT-1 gene in transgenic plants was significantly increased.【Conclusion】SsREMO-1a can improve the drought resistance of plants and regulate CAT-1 gene to some extent.Download(CAJ format)Download(PDF format)

关 键 词：割手密 REMO基因 干旱胁迫 定量表达 功能分析 

分 类 号：S566.1[农业科学—作物学]