Abstract: To resolve the partial compensation effect of the different srGAPs in neuronal development, a miR-shRNA expression system was constructed which can down regulate srGAP family members srGAP1—3 simultaneously. Multiple miRshRNA oligos targeting different srGAPs were designed, synthesized and then inserted into the miRNA expression vector to generate miRshRNA against srGAP1—3 respectively. Western blot was performed to detect the knockdown effect of different miR-shRNAs, the three miRshRNA with the most obvious knockdown effect to srGAP1—3 respectively were selected. Then the miR-shRNAs were chained together in one vector to generate a chaining miRshRNA expression vector. The results show that the effective miRshRNA against corresponding srGAP1—3 are generated and named J17, J24 and J33. Furthermore, the chaining miR-shRNA J1+2+3 against srGAP1—3 is obtained.

Key words: Slit-Robo GTPase-activating protein (srGAP), short hairpin RNA (shRNA), RNA interference, Western blot