摘要
利用PCR技术扩增目的基因FGF13VY亚型,经酶切、连接等反应将目的基因插入穿梭载体p Shuttle-CMV中,进而转化至含有Ad Easy质粒的大肠杆菌中,进行重组。将所产生的腺病毒载体转染在HEK-293细胞中,进行腺病毒的包装、分泌和扩增,将所获得的腺病毒在背根神经节中进行过表达,可检测到腺病毒携带的绿色荧光蛋白表达在背根神经节神经元,q PCR检测到FGF13VY mRNA的表达明显升高。FGF13VY腺病毒过表达载体的成功构建,为研究其在DRG神经元中作用奠定了基础。
FGF13VY gene was amplified by PCR technique and inserted into pShuttle-CMV vector by enzyme digestion and ligation reaction.Then FGF13VY-pShuttle-CMV was transformed into E.coli containing AdEasy plasmid for recombination.The produced adenovirus vector was transfected into HEK-293 cells to package and amplify adenovirus,then the adenovirus was overexpressed in DRG neurons.Green fluorescent protein carried by adenovirus can be detected in DRG neurons.The expression of FGF13VY mRNA was significantly increased by qPCR.The successfully constructed FGF13VY overexpression adenovirus vector lays an experimental foundation for study of the role of FGF13VY in DRG neurons.
作者
董梅
陈宇峰
孙如梦
华乐
杨甫
孟丹琳
杨晶
王川
Dong Mei;Chen Yufeng;Sun Rumeng;HUA Le;YANFG Pu;MENG Danlin;YANG Jing;WANG Chuan(The Affiliated Hospital of Hebei University of Science and Technology,Shijiazhuang 050018;College of Basic Medicine,Hebei Medical University,Shijiazhuang 050017)
出处
《安徽医科大学学报》
CAS
北大核心
2018年第12期1956-1959,共4页
Acta Universitatis Medicinalis Anhui
基金
国家自然科学基金(编号:81770407)
河北省自然科学基金(编号:H2017206262
C2018206277)
河北省高等学校科学技术研究项目(编号:ZD2015007
ZD2016002
Z2015005)
河北省高层次人才资助项目(编号:A2017005070)
河北医科大学大学生创新性实验计划项目(编号:USIP201509A)
