摘要
【目的】克隆马铃薯茄啶鼠李糖基转移酶基因(sgt3)的启动子序列并对其进行功能鉴定.【方法】采用染色体步移技术(genome walking)进行启动子的克隆,构建了该启动子驱动报告基因gfp::gus的植物双元表达载体p1304sgt3p,以pCAMBIA1304为阳性对照表达载体,采用农杆菌介导的烟草叶片瞬时表达及稳定遗传转化,结合GUS组织化学染色对其进行功能鉴定.【结果】电泳图表明为1条长约2 500bp的特异扩增条带,瞬时表达和稳定遗传表达的烟草叶片的GUS组织化学染色结果均显示gus基因在转化烟草叶片中高效表达,并且低于阳性对照,非转化烟草叶片中无表达.【结论】成功克隆到sgt3基因起始密码子上游2392bp的启动子序列并且该启动子具有活性.
【Objective】To clone the promoter sequence of 5′upstream of initial codon of the potato gene of solanidine galactosyltransferases（sgt3）and identify its function.【Method】The promoter sequence was cloned by using the Genome walking and then was fused with gfp：：gus gene originated from pCAMBIA1304 to construct a binary vector p1304sgt3 p.The vector pCAMBIA1304 was used as a positive control expression vector.And they were introduced into wild type tobacco leaves through Arobacterium mediated instantaneous expression and stable genetic transformation to identify the function of the promoter sequence.【Result】Electrophoregram shows that a pecific amplification bands about 2 500 bp was cloned.The GUS staining showed that gus gene was highly expressed both in transgenic tobacco leaves and instantaneous expression blade,and the expression was lower than that in positive control,and there was no epression in the wild type tobacco leaves.【Conclusion】The 2 392 bp promoter sequence of 5′upstream of initial co-don of sgt3 gene cloned was successfully cloned,and the promoter had activity.
出处
《甘肃农业大学学报》
CAS
CSCD
北大核心
2016年第5期32-38,共7页
Journal of Gansu Agricultural University
基金
国家自然科学基金项目(31260343)
科技部国家重点基础研究发展计划(973计划)前期项目(2010CB13400)
甘肃省自然科学基金项目(0710RJZA088)
关键词
马铃薯
茄啶鼠李糖基转移酶
启动子克隆
功能鉴定
solanum tuberosum
solanidine rhamnosyltransferase
promoter cloning
functional identification
