Construction and identification of a eukaryotic expression vector containing human homeobox gene.
- Author:
Tian-Lan LI
1
;
Chun-Ting ZHAO
;
Zhong-Guang ZHANG
;
Zhu-Zhen LIU
;
Shi-Hai LIU
;
Dong-Mei MENG
;
Yuan-Feng HANG
Author Information
1. Department of Hematology, Qingdao University Medical College Affiliated Hospital, Qingdao 266003, Shangdong Province, China.
- Publication Type:Journal Article
- MeSH:
Cell Line;
Cloning, Molecular;
Gene Expression;
Genes, Homeobox;
Genetic Vectors;
Humans;
Plasmids;
Transfection
- From:
Journal of Experimental Hematology
2011;19(3):721-724
- CountryChina
- Language:Chinese
-
Abstract:
In order to investigate the special role of HOXB4 in expansion and self renewal of hematopoietic stem cells, the cDNA of HOXB4 was extracted and cloned from umbilical cord blood mononuclear cells by using RT-PCR. Then the eukaryotic expression bicistronic plasmid vector pIRES2-EGFP/HOXB4 was designed and constructed after cutting HOXB4 and pIRES2-EGFP respectively by restriction enzyme EcoRI and BamHI. The recombinant plasmid was delivered into competent cells of Escherichia coli. The successful construction of plasmid was confirmed by the identification of endonuclease cutting and sequencing. The results showed that the HOXB4 cDNA was cloned successfully from umbilical cord blood mononuclear cells and the recombinant eukaryotic expression bicistronic plasmid vector was constructed, and then introduced it into 293T cells successfully. It is concluded that a pIRES2-EGFP/HoxB1 eukaryotic expression bicistronic plasmid vector has been constructed successfully, which results provide a useful material basis for exploration of HoxB4 function in the proliferation and differentiation of hematopoietic cells.
