Establishment of a colorectal cancer cell line with PRL-3 and CDH22 gene knock-down by lentivirus-mediated RNA interference / 南方医科大学学报

ABSTRACT

<p><b>OBJECTIVE</b>To 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.</p><p><b>METHODS</b>A 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.</p><p><b>CONCLUSIONS</b>A human colorectal cancer cell line with stable PRL-3 and CDH22 gene knock-down has been established.</p>