Construction and identification of lentiviral vector of siRNA specific for Beclin1 gene / 生物医学工程学杂志
Journal of Biomedical Engineering
; (6): 131-135, 2013.
Article
in Zh
| WPRIM
| ID: wpr-234690
Responsible library:
WPRO
ABSTRACT
The lentiviral vector was used for construction of a recombinant mediating RNA interference (RNAi) against Beclin1 gene in this study. Recombinant vector plasmid was transfected into non small cell lung cancer (NSCLC) A549 cells by liposome. PCR results showed that three amplified positive fragments were inserted into pRNAT-U6. 2/Lenti vectors. DNA sequencing results showed that the three recombinant lentivirus plasmids, pRNAT-U6. 2/Lenti-si356, pRNAT-U6. 2/Lenti-si423 and pRNAT-U6. 2/ Lenti-si684 were constructed successfully. After transfection with liposome, RT-PCR and Western blot analysis confirmed that the expression of Beclin1 mRNA and protein was inhibited in the three recombinant lentivirus plasmids transfected groups, and gene silencing efficacy was 35.56%, 89.22% and 66.78%, respectively. The results demonstrated that the lentiviral vectors of RNAi targeting Beclin1 gene were successfully constructed, and NSCLC A549 stable cell line with Beclin1 gene knockdown was established. This study finally provided a new cell model to explore the biological behavior of the Beclin1 gene in NSCLC A549 cells.
Full text:
1
Index:
WPRIM
Main subject:
Pathology
/
Autophagy
/
Molecular Sequence Data
/
Base Sequence
/
Transfection
/
Carcinoma, Non-Small-Cell Lung
/
Lentivirus
/
RNA, Small Interfering
/
RNA Interference
/
Cell Line, Tumor
Type of study:
Prognostic_studies
Limits:
Humans
Language:
Zh
Journal:
Journal of Biomedical Engineering
Year:
2013
Type:
Article