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Meanwhile, plasmid pYES2/CT (Invitrogen, Carlsbad, CA, United States) was first digested with Eco RI and Xba I, gel-purified, and ligated with the above two products using ClonExpress Ultra One Step Cloning Kit (Vazyme, Nanjing, China) and then transformed into Escherichia coli DH5a cells following the manufacturers instructions