Objective: To construct prokaryotic expression recombination plasmids containing GST-GSK3β1,GST-GSK3β2 and dominant-negative mutant GST-GSK3β1 K85M,then express,purify and identify these three fusion proteins. Methods: Using PCR and recombinant DNA technology,the target genes were amplified from templates Flag-GSK3β1,YFP-GSK3β2 and Flag-GSK3β1 K85M respectively and inserted into the prokaryotic expression vector pGEX-6P-1 with glutathione-S-transferase(GST) tag to construct the expression plasmids expressing GST-GSK3β1,GST-GSK3β2 and GST-GSK3β1 K85M fusion proteins. An effective mean of expression and purification was established by a series of expression,gentle lysis and purification experiments. In the end,Western blot was used to identify the purified fusion protein. Results: We succeed in constructing wild type and dominant-negative mutant plasmids of GST-GSK3β,expressing and purifying these fusion proteins. Conclusion: The successful construction,expression and purification of those target proteins facilitate us to further study the biology function of GSK3β.