大鼠BDNF基因克隆和重组逆转录病毒pLXSN-BDNF的构建  

Cloning rat BDNF gene and construction of a recombinant retroviral vector pLXSN-BDNF

作　　者：邵正波[1] 原慧萍[1] 周欣荣[1] 李鸿翼[1] 曲巍[1] 杨滨滨[1] 

机构地区：[1]哈尔滨医科大学附属第二医院眼科,黑龙江省哈尔滨市150086

出　　处：《眼科新进展》2006年第11期814-817,共4页Recent Advances in Ophthalmology

基　　金：国家自然科学基金资助(编号:30471844);哈尔滨医科大学研究生创新基金资助~~

摘　　要：目的克隆大鼠脑源性神经营养因子(brain-derived neurotrophic factors,BDNF)基因并构建含BDNF基因片段的真核表达载体pLXSN-BDNF,用于视神经损伤修复的基因治疗。方法利用RT-PCR的方法从大鼠海马组织的总RNA中扩增BDNF基因片段,序列两端分别引入限制性内切酶识别位点EcoR I和Xho I,并将其定向克隆入pMD18-T Si mple载体,测序正确后亚克隆入逆转录病毒载体pLXSN。结果经过PCR、酶切和测序鉴定,所克隆的BDNF基因序列正确,克隆的BDNF基因已经正确插入到逆转录病毒载体pLXSN中。结论成功克隆了BDNF基因并且构建了真核表达载体pLXSN-BDNF,为进一步研究基因治疗视神经损伤疾病奠定基础。Objective To clone the rat brain-derived neurotrophic factors （BDNF） gene and develop a modified retroviral vector for the protection of optic nerve injury. Methods The cDNA encoding the rat BDNF gene was amplified by RT-PCR from total RNA of the rat hippocampal tissue and the restriction sites EcoR I and Xho I were introduced into the ends of BDNF cDNA. Being verified with DNA sequencing, the BDNF gene was cloned into pMD18-T Simple vector, and then subcloned into the plasmid pLXSN. Results With PCR, restriction analysis and DNA sequencing, the sequence of the BDNF gene was correct and the clone gene was inserted into the retroviral vector exactly. Conclusion The BDNF gene has been cloned correctly and the recombinant retroviral vector carrying BDNF gene has been constructed successfully, and all these lay foundations for a further study of gene therapy for the retinal diseases.

关 键 词：脑源性神经营养因子 逆转录病毒载体 基因治疗 

分 类 号：R373[医药卫生—病原生物学]