Construction and identification of the recombinant eucaryotic plasmid PcDNA3.1-MxA
- VernacularTitle:人MxA基因重组真核载体的构建及鉴定
- Author:
Qin HUANG
- Publication Type:Journal Article
- Keywords:
MxA;
PcDNA3.1;
PcDNA3.1-MxA
- From:
Journal of Chongqing Medical University
1986;0(02):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To construct and identify the recombinant eukaryotic plasmid PcDNA3.1-MxA encoding MxA with the eukaryotic plasmid PcDNA3.1.Methods:Recombinant plasmid PAdTrack-MxA constructed before and PcDNA3.1 were amplified in Escherchia coli JM109.Plasmids were double-digested with restrictive endonucleases NotⅠ and XbaⅠ after extracted,and then ligated to construct recombinant eukaryotic plasmid PcDNA3.1-MxA.The recombinant plasmid was selected for Ampicillin resistance and then confirmed by digestion with NotⅠ and XbaⅠ,PCR and sequencing.The sequence was homology-analysed compared with the sequence of MxA gene in PAdTrack-MxA and the sequence of MxA gene published in Genebank(NM_002462) with sofeware DNAssist 2.0.Results:The restrictive endonuclease and PCR analysis confirmed that the recombinant eukaryotic plasmid PcDNA3.1-MxA was constructed successfully.Sequencing analysis revealed that the cloned segment was 2012bp including MxA gene sequence,and the nucleotide sequence of MxA gene was same to that in PAdTrack-MxA exactly.There were five variations in nucleotide sequence compared with published sequence in Genebank,which caused only one amino acid residue replaced from isoleucine to valine.This replaced amino acid residue was located in the non functional area of MxA protein.Conclusions:The recombinant eukaryotic plasmid PcDNA3.1-MxA is constructed successfully,which lays foundation for the further study on anti-HBV effects of MxA.
