中文摘要：

目的建立稳定干扰PRL-3及CDH-22基因的结直肠癌细胞株，为探讨PRL-3、CDH22及其相互作用在结直肠发生及转移中的作用提供理想的细胞模型。方法以稳定干扰人PRL-3基因大肠癌SW480细胞的克隆为基础，以靶向人CDH22基因的RNAi慢病毒再次感染该细胞株，将经慢病毒二次感染的细胞悬液通过有限稀释法制备细胞单克隆，各细胞克隆扩大培养，荧光定量PCR检测各细胞克隆PRL-3及CDH22mRNA表达水平。结果应用于二次感染的慢病毒悬液的滴度为8×10^5U／ml。综合分析荧光定量PCR结果，所获得的细胞克隆中1号克隆PRL-3及CDH22mRNA水平的表达显著降低。结论成功建立PRL-3及CDH-22基因稳定敲低的大肠癌细胞克隆，探索同时敲低2个基因的慢病毒RNAi技术。

英文摘要：

Objective 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 eolorectal cancer. Methods 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 ViraPowerTM 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 ×10^5 U/ml. Seventeen positive clones were selected, among which the Clone 1 exhibited substantially down-regulated CDH22 and PRL-3 mRNA expressions. Conclusion A human coloreetal cancer cell line with stable PRL-3 and CDH22 gene knock-down has been established.