[Construction and Identification of Leukemia Cell Line Stably Expressing CD123 and CLL-1].
Zhongguo Shi Yan Xue Ye Xue Za Zhi
; 29(2): 322-327, 2021 Apr.
Article
em Zh
| MEDLINE
| ID: mdl-33812394
ABSTRACT
OBJECTIVE:
To construct an acute myeloid leukemia cell line stably expressing CD123-CLL1 so as to provide an "in vitro" model for studying the role of CD123 and CLL-1 in leukemia and the treatment targeting CD123 and CLL-1.METHODS:
The recombinant plasmid of lentivirus was constructed by synthesizing CD123 and CLL-1 sequences and PCR homologous recombination. The lentivirus vector was packaged by three-plasmid packaging system. After collecting the supernatant of lentivirus, the virus titer was determined by quantitative PCR. K562 leukemia cells were collected and transtected with virus supernatant. Leukemia cell line stably expressing the target gene were screened by purinomycin. The expression levels of CD123 and CLL-1 were detected by RT-PCR and flow cytometry.RESULTS:
The lentiviral vector was successfully constructed, and identified by agarose gel electrophoresis and gene sequencing, then the virus titer of the supernatant was up to 5.81×108 after quantitative PCR assay. The K562 leukemia cell line obtained positive expression cells after being infected by puromycin. The high expression of CD123 and CLL-1 was confirmed by RT-PCR, while the significantly high expression of CD123 and CLL-1 was confirmed by flow cytometry.CONCLUSION:
Lentiviral vector expressing CD123-CLL1 has been successfully constructed, and K562 leukemia cell line stably expressing CD123 and CLL-1 has been successfully obtained.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Leucemia Linfocítica Crônica de Células B
/
Subunidade alfa de Receptor de Interleucina-3
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
Zh
Revista:
Zhongguo Shi Yan Xue Ye Xue Za Zhi
Assunto da revista:
HEMATOLOGIA
Ano de publicação:
2021
Tipo de documento:
Article
País de afiliação:
China