Abstract
Pancreatic cancer is one of the most deadly cancers in the world and metastasis is often the most common cause of cancer death. While many recent studies have used the micro-array strategy to identify various metastasis-related genes, we suspect that many others still remain to be discovered. Our goal is to use a function-based robust shRNA screening strategy that may complement micro-array methods in identifying novel metastasis-related genes. There are several pooled shRNA sub-libraries available for screening. As an initial attempt, we planed to use the pooled protein kinases/phosphatases shRNA clones for this purpose, because they directly regulate protein phosphorylation and affect down-stream signaling. First, we examined the migration and invasion abilities of four pancreatic cell lines AsPc-1, BxPc-3, SUIT-2 and PANC-1 and determined to use SUIT-2 cells which have the lowest invasiveness potential for future study. We also tried to optimize the infection efficiency of the shRNA lentiviruses by testing the cell seeding density, puromycin sensitivity and virus tittering before infect these cancer cells with a pooled shRNA lentiviruses. We established pure clones from these invading cells and determined the identity of each individual shRNA clones by sequencing the PCR products after the shRNA fragments were amplified. Finally we tried to identify the candidate invasiveness-associated genes targeted by the corresponding shRNA clones, and to investigate the biological functions, hoping that the newly identified genes will potentially serve as targets for novel cancer therapies development.