Establishment of a colorectal cancer cell line with PRL-3 and CDH22 gene knock-down by lentivirus-mediated RNA interference.
- Author:
Juan YAO
1
;
Yan-qing DING
;
Jun ZHOU
;
Yu-hong LIU
;
Jian-ming LI
Author Information
- Publication Type:Journal Article
- MeSH: Cadherins; genetics; Cell Line, Tumor; Clone Cells; metabolism; Colorectal Neoplasms; genetics; pathology; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; methods; Humans; Lentivirus; genetics; Neoplasm Proteins; genetics; Protein Tyrosine Phosphatases; genetics; RNA Interference; RNA, Messenger; genetics; metabolism
- From: Journal of Southern Medical University 2009;29(4):593-597
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo establish a colorectal cancer cell line with stable PRL-3 and CDH22 gene knock-down for investigating the role of PRL-3 and CDH22 genes in the carcinogenesis and metastasis of colorectal cancer.
METHODSA recombinant lentiviral vector targeting CDH22 gene was obtained using the pENTRTM/U6 construct and pLenti6/BLOCK-iT (TM)-DEST vector. The recombinant lentivirus was harvested from 293FT cells cotransfected with the optimized ViraPower(TM) Packaging Mix and the pLenti6/BLOCK-iT(TM)-DEST expression construct. SW480/PRL-3- cells were infected with the recombinant lentivirus targeting CDH22, and SW480 cells with stable PRL-3 and CDH22 knock-down were screened by blasticidin selection. PRL-3 expression in the cells was determined by real-time RT-PCR. RESULTS The titer of the lentivirus for the second infection was 8 x 10(5) U/ml. Seventeen positive clones were selected, among which the Clone 1 exhibited substantially down-regulated CDH22 and PRL-3 mRNA expressions.
CONCLUSIONSA human colorectal cancer cell line with stable PRL-3 and CDH22 gene knock-down has been established.
