[Construction and identification of rhesus monkey Schwann cells modified with human glial cell derived neurotrophic factor gene].
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi
; 27(11): 1363-7, 2013 Nov.
Article
em Zh
| MEDLINE
| ID: mdl-24501898
ABSTRACT
OBJECTIVE:
To construct the rhesus monkey Schwann cells (SCs) modified with human glial cell derived neurotrophic factor (hGDNF) gene.METHODS:
The coding sequence ofhGDNF amplified by PCR from pUC19-hGDNF was inserted into eukaryotic expression vector pBABE-puro. The recombinant eukaryotic expression vector pBABE-puro-hGDNF was identified with restriction enzyme digestion and DNA sequencing. The SCs were isolated from rhesus monkeys, cultured and purified. The SCs were transfected with the recombinant retrovirus vector containing hGDNF gene. The mRNA and protein expressions of hGDNF were analyzed by real-time fluorescent quantitative PCR and Western blot.RESULTS:
The PCR product of hGDNF coding sequence was a 596 bp specific segment. The recombinant eukaryotic expression vector was digested into a 596 bp specific segment by specific restriction enzyme and another segment. The 596 bp segment confirmed by DNA sequencing was consistent with hGDNF sequence on GenBank. Restriction enzyme digestion and sequencing results showed that the coding sequence of hGDNF was successfully inserted into the recombinant retrovirus vector and the mRNA and protein expressions of hGDNF were significantly higher in transfected SCs than non-transfected SCs (P < 0.05).CONCLUSION:
The rhesus monkey SCs modified with hGDNF gene are successfully constructed and hGDNF can be released continuously and stably, which will provide a foundation for the further research about cell therapy of hGDNF-SCs in the repair of injured nerve.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Retroviridae
/
Células de Schwann
/
Fator Neurotrófico Derivado de Linhagem de Célula Glial
/
Vetores Genéticos
Tipo de estudo:
Diagnostic_studies
/
Prognostic_studies
Limite:
Animals
/
Humans
/
Male
Idioma:
Zh
Revista:
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi
Ano de publicação:
2013
Tipo de documento:
Article