Construction of vector of recombinant adeno-associated virus including NT4-GFP-Ant fusional reporting gene and significance
- VernacularTitle:携带信号肽NT4-GFP-穿膜肽Ant融合基因的重组腺相关病毒载体的构建及意义
- Author:
Hao WU
;
Jiang WU
;
Yu YANG
;
Xin SUN
;
Guangxiao YANG
;
Quanying WANG
- Publication Type:Journal Article
- Keywords:
green fluorescent protein;
neurotrohphin-4;
antennapedia;
gene clone
- From:
Journal of Jilin University(Medicine Edition)
2006;0(05):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct NT4-GFP-Ant fusional reporting gene and the vector of NT4-GFP-Ant recombinant adeno-associated virus(AAV).Methods The GFP gene was cloned by using PCR and T-vector cloning method.The positive clone was identified by the restriction enzymes,and then the cloned amplified fragments were sequenced and analyzed.The resulting gene of GFP and Ant,PBV220/NT4 were connected by DNA ligase,and thus PBV220/NT4-GFP-Ant was constructed,then the NT4-GFP-Ant fragment was gained and identified by the restriction enzymes.The resulting gene of NT4-GFP-Ant fragment was inserted into the EcoRⅠ-BamHⅠsite of vector plasmid pSSHG to construct the vector of NT4-GFP-Ant recombinant AAV.Results A 730 bp fragment of DNA was gained when T-easy/GFP was cut by the restriction enzyme EcoRⅠ.The cloned GFP gene was coincident with the sequence in GenBank.A 1 000 bp fragment of DNA was gained when pBV220/NT4-GFP-Ant was cut by the restriction enzymes BamHⅠ/EcoRⅠ.A 1 000 bp fragment of DNA was gained when PSSHG/NT4-GFP-Ant was cut by the restriction enzymes BamHⅠ/EcoRⅠ.Conclusion GFP gene is cloned successfully,NT4-GFP-Ant gene and PSSHG/NT4-GFP-Ant recombinant AAV vector are constructed successfully.
