Abstract |
The enhanced green fluorescent protein (EGFP) pEGFP-N1-P53 eukaryotic expression vector, which contains the human tumor suppressor p53, was constructed and transfected into chicken fibroblast cells and stage-X blastoderm to analyze the transfection efficiency. The complementary DNA of the human p53 gene was cloned by reverse transcription-polymerase chain reaction from human peripheral blood and inserted into the pEGFP-N1 vector by HindIII and BamHI double digestion. The pEGFP-N1-P53 vector was transfected into chicken embryo fibroblasts by Lipofectamine 2000 liposomes, and the transfection efficiency was analyzed by fluorescence microscope after 36 h of transfection. The stage-X blastoderm was also transfected by blastoderm injection using Lipofectamine 2000 liposomes at room temperature after 12-24 h; then hatching occurred until seventh day, and the transfection efficiency was analyzed by fluorescence microscope in the dead embryo. A total of 90 hatching eggs were transfected by the pEGFP-N1-P53 vector, and 20 chicken embryos expressed the reporter gene, which indicated that recombinant pEGFP-N1-P53 could be transfected and expressed in stage-X blastoderm by liposomes. Chicken embryo fibroblasts were transfected and expressed the reporter gene. The pEGFP-N1-P53 vector was constructed successfully and could be transfected and expressed in chicken embryo fibroblasts and stage-X blastoderms efficiently.
|
Authors | Z Song, Z H Li, X Q Lei, T S Xu, X H Zhang, Y J Li, G M Zhang, S M Xi, Y B Yang, Z G Wei |
Journal | Genetics and molecular research : GMR
(Genet Mol Res)
Vol. 14
Issue 1
Pg. 931-9
(Feb 02 2015)
ISSN: 1676-5680 [Electronic] Brazil |
PMID | 25730031
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
|
Chemical References |
- Recombinant Fusion Proteins
- TP53 protein, human
- Tumor Suppressor Protein p53
- enhanced green fluorescent protein
- Green Fluorescent Proteins
|
Topics |
- Animals
- Blastoderm
(growth & development, metabolism)
- Chick Embryo
- Fibroblasts
(metabolism)
- Gene Expression Regulation, Developmental
- Genetic Vectors
- Green Fluorescent Proteins
(genetics)
- Humans
- Recombinant Fusion Proteins
(biosynthesis, genetics)
- Tumor Suppressor Protein p53
(biosynthesis, genetics)
|