[Construction and identification of recombinant adenovirus vector containing Tet-on adjustable NT3 and BDNF genes].
OBJECTIVE
To construct Tetracycline(tet) inducible recombinant adenovirus vectors containing human neurotrophin 3(NT3) and brain-derived neurotrophic factor(BDNF) genes, respectively and perform PCR and restrictive enzyme digestion analysis.


METHODS
Full length NT3 and BDNF cDNAs were subcloned into pIND vector, followed by being cloned into pTRE-shuttle2 vector. The NT3 and BDNF gene fragments resulted from the pTRE-shuttle2-NT3 and pTRE-shuttle2-BDNF digested with I-Ceu I and PI-Sce I were linked to the linear adeno-X virus DNA. The recombinant adenovirus vectors were confirmed by PCR and restriction enzyme digestion analysis.


RESULTS
The PCR identification showed that a given band with 312 bp, and predictive fragments proved by restriction enzyme digestion analysis were exhibited. All the above results indicated that human NT3 and BDNF genes had been connected with pAdeno-X vectors correctly.


CONCLUSIONS
Tet inducible recombinant adenovirus vector of Human NT3 and BDNF genes have been constructed successfully, which upon packaged in HEK293 cells, will be used to introduce the target genes into Schwann's cells in-vitro or in-vivo.
Li
Zhi-quan
ZQ
The Institute of Orthopaedics, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, China.
Zhu
Qing-sheng
QS
Zhu
Jin-yu
JY
Hao
Wei
W
Xu
Yan-ming
YM
Wang
Cheng-ji
CJ
Yang
An-gang
AG
D000256:Adenoviridae / Q000235:genetics*; D019208:Brain-Derived Neurotrophic Factor / Q000096:biosynthesis / Q000235:genetics*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D018274:Electroporation; D004622:Embryo, Mammalian; D005822:Genetic Vectors; D006801:Humans; D007668:Kidney / Q000166:cytology / Q000378:metabolism*; D020933:Neurotrophin 3 / Q000096:biosynthesis / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011995:Recombination, Genetic; D015183:Restriction Mapping; D013752:Tetracycline / Q000494:pharmacology*
D019208:Brain-Derived Neurotrophic Factor; D020933:Neurotrophin 3; D013752:Tetracycline