Construction and transformation of yeast bait plasmids carrying synapse growth factor Neuritin cDNA fragment

  Objective   To construct and transform yeast bait plasmid carrying the synapse growth factor neurit in cDNA fragment.

  Methods   Fragment of ORF of full neuritin cDNA was amplified using PCR and directly ligated to the pLex Avector.Insert-contained plasmid was confir med by restriction analysis and DNAsequencing and then transformed into EGY 48 p8op-LacZyeast strain.

  Results   Recombinant pLexA-neuritin plasmid was successfully constrcted.Two sort of yeasts respectively transformed recombinant plasmids and empty pLex A vector could grow white colonies on SD/Gal/Raf/-His/-Ura plates(while the yeasts transformed pLexA-pos, positive control plasmid were blue colonies)and none could survive on SD/His/-Leu/-Ura plates.After being cultured in SD/-His/-Ura liquid medium for 16 h, the A6 00 of them were both 0.9 respectively.This indicated that the fusion proteins expressed by recombinant plasmids did not activate the expression of yeast reporter genes LEU 2 and lacZ, and had no toxicity to yeast strain.

  Conclusion   The bait plasmids constructed can be used to study the funct ion of neuritin in Yeast Two-Hybrid Screen.