Expression of an angiogenesis inhibitor METH1 and assay of activation of reporter genes in yeast double-hybrid system

Objective This work is intended as a basis for further study of the molecular mechanism of METH1 inhibition of hypertrophic scar, through the expression of cDNA fragments of METH1 and assay of activation of reporter genes in yeast double-hybrid system. Methods cDNA fragments of METH1 were amplified by the polymerase chain reaction (PCR), and were subsequently cloned into pUC19. After verification by sequencing, they were subcloned into the vector pGBKT7 of the yeast double-hybrid system. These recombinant plasmids were then transfered into yeast fungus AH109, and their expression products were assayed for ability to activate the reporter genes. Results The cDNA fragments of METH1 were amplified successfully. The expression product of fragments were not toxic to AH109 and could not activate the reporter genes. Conclusion The expression product of the active fragment of METH1 can be used as a to search for interacting proteins in this yeast double-hybrid system.