[Comparison of three methods in construction fusion gene of influenza A virus Nucleoprotein].
OBJECTIVE
To develop a best method of constructing influenza NP fusion gene containing enhanced green fluorescent protein (EGFP).


METHODS
The full-length NP gene of influenza A was amplified by RT-PCR and was inserted into an eukaryotic expression vector pEGFP-N1 in order to construct a fusion gene of pEGFP-N1-NP using three different methods. Method one, NP gene containing restriction endonucleases and pEGFP-N1 were both digested using the same restrict enzymes and ligated, yielding the fusion gene of pEGFP-N1-NP. Method two, NP gene was cloned into pMD19-T Vector to construct a plasmid of pMD19-T-NP. The pMD19-T-NP cloned into pEGFP-N1 to construct the fusion gene of pEGFP-N1-NP; Method three, NP gene containing restriction endonucleases was cloned into pMD19-T Simple Vector to construct a plasmid of pMD19-T-NP. The pMD19-T-NP cloned into pEGFP-N1 to construct the fusion gene of pEGFP-N1-NP.


RESULTS
The fusion gene of recombinant eukaryotic expression vector pEGFP-N1-NP was successfully constructed by using method three.


CONCLUSIONS
The full-length NP gene is obtained and its fusion gene of recombinant eukaryotic expression plasmid is successfully constructed. This study provides foundation for further understanding the biological function of NP protein and the mechanism of diseases induced by influenza A virus.
Department of Respiratory Medicine, Laboratory of Respiratory Medicine Research, Affiliated Hospital of Zunyi Medical College, Zunyi 563003, China.
Bo
B
Liu
Bao-Lin
BL
Wang
Ling
L
Chen
Jing
J
Yan
Jian-Yong
JY
Zhang
Hong-Xuan
HX
He
Hong
H
Zhang
D019677:Artificial Gene Fusion; D003001:Cloning, Molecular; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics*; D019590:Nucleocapsid Proteins; D016133:Polymerase Chain Reaction; D016601:RNA-Binding Proteins / Q000235:genetics*; D014758:Viral Core Proteins / Q000235:genetics*
C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; D016601:RNA-Binding Proteins; D014758:Viral Core Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins