【作者】 司爱君； 陈红； 余渝； 孔宪辉； 刘丽； 王旭文； 田琴；

【Author】 Si Aijun;Chen Hong;Yu Yu;Kong Xianhui;Liu Li;Wang Xuwen;Tian Qin;Cotton Research Institute,Xinjiang Academy of Agricultural and Reclamation Sciences/Key Laboratory of China Northwestern Inland Region,Ministry of Agriculture/Key Laboratory of Genetic Improvement and High Yield,Xinjiang Production and Construction Corps;

【机构】 新疆农垦科学院棉花研究所/农业部西北内陆区棉花生物学与遗传育种重点实验室/新疆兵团棉花遗传改良与高产栽培重点实验室；

【摘要】 通过反转录聚合酶链式反应(Reverse transcription-polymerase chain reaction,RT-PCR)筛选到1个高表达的棉花叶片特异性表达基因。以陆地棉Coker 312基因组DNA为模板,通过染色体步移法(Genome walking)克隆获得翻译起始位点上游2 kb左右的DNA片段,将其命名为叶片特异性表达启动子(Leaf specific promoter,LSP)。PlantCARE在线分析表明,GhLsp启动子序列除了含有核心元件及顺式元件TATA-box、CAAT-box外,还存在光反应元件、低温诱导元件、MYB干旱诱导结合位点等多个特异性作用元件。通过构建该启动子驱动GUS的植物表达载体pGhlsp::GUS,并经农杆菌介导的花序侵染法转化拟南芥,对转基因拟南芥进行GU S组织化学染色观察,结果显示该基因主要在叶片中特异性表达,而在根部以及茎部几乎不表达。作为起重要调控作用的光诱导启动子,这类启动子将在抗虫、抗病、抗旱、抗高渗胁迫等抗逆性的转基因植物优良品种选育等方面应用广泛,可实现外源基因表达的定时、定点、定量调控。此类启动子用于基因工程,可以在改良棉花性状的同时减少对棉花生理方面的副作用,在棉花抗逆转基因育种方面有广泛的应用前景。更多还原

【Abstract】 According to reverse transcription-polymerase chain reaction(RT-PCR),a leaf-specific expression gene of a high expression level was discovered and verified in cotton,a DNA fragment of about 2 kb of the upstream promoter sequence of the gene was successfully obtained by the Genome Walking method using Gossypium hirsutum cv.Coker 312 genomic DNA as a template,and named leaf specific promoter(LSP).PlantCARE online analysis showed that not only the core elements TATA-box and CAAT-box but also a plurality of homeopathic action elements of the promoter were present in GhLsp promoter sequence.To further understand its function,the plant expression vector of promoter-driven GUS pGhlsp::GUS was constructed and transformed into Arabidopsis mediated by Agrobacterium tumefaciens infection.The expression pattern of the promoter was analyzed by GUS staining.The results showed that the gene was mainly expressed in leaves,while almost non-expressed in the roots and stems.As a light-inducible promoter that plays an important regulatory role,this kind of promoter will be widely used in the breeding of transgenic plants with excellent resistance to insects,disease resistance,drought resistance and hypertonic stress,to achieve exquisite gene expression and control with the fixed time,fixed location,fixed quantitative.The use of these promoters in genetic engineering can improve the cotton traits while reduce the physiological side effects to cotton,so it has the wide spread application prospect in anti-breeding.更多还原