Construction and Identification of Leukemia Cell Line Stably Expressing CD123 and CLL-1.
10.19746/j.cnki.issn.1009-2137.2021.02.003
- Author:
Xiang-Yu WANG
1
;
Guo-Qiang LIN
1
;
Lei YU
2
;
Li-Qing KANG
2
;
Jing-Wen TAN
2
;
Yan-Ming ZHANG
3
;
De-Pei WU
4
Author Information
1. Department of Hematology, Huai'an Hospital Affiliated to Xuzhou Medical College and Huai'an Second People's Hospital, Huai'an 223002, Jiangsu Province, China.
2. Shanghai Unicar-Therapy Biomed-Phamaceutical Technology Co., Shanghai 201203, China.
3. Department of Hematology, Huai'an Hospital Affiliated to Xuzhou Medical College and Huai'an Second People's Hospital, Huai'an 223002, Jiangsu Province, China,E-mail: zhangyanming2005@126.com.
4. Department of Hematology, The First Affiliated Hospital of Soochow University, Suzhou 215006, Jiangsu Province, China,E-mail: wudepei@suda.edu.cn.
- Publication Type:Journal Article
- MeSH:
Cell Line, Tumor;
Genetic Vectors;
Humans;
Interleukin-3 Receptor alpha Subunit;
K562 Cells;
Lentivirus/genetics*;
Leukemia, Lymphocytic, Chronic, B-Cell/genetics*;
Plasmids;
Transfection
- From:
Journal of Experimental Hematology
2021;29(2):322-327
- CountryChina
- Language:Chinese
-
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 5.81×10
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.
