To mutate selected amino acids in the N-domain of PSG1, we designed a PSG1 cDNA containing an Acc65I and an XhoI restriction site recognition sequence as silent mutations and cloned the cDNA into pFuse-IgG1 e3-Fc1 vector (InvivoGen). A 406-bp cDNA fragment was synthesized by Genscript Corporation in which the nucleotides coding for amino acids G and D-in positions 93 and 95, respectively, of the N-domain of the mature PSG1-were replaced for nucleotides coding for amino acids S and L, respectively. The fragment containing the mutations had an EcoRI site at the 5 0 end and an Acc65I site at the 3 0 end. To generate the mutant, we replaced the EcoRI-Acc65I fragment in PSG1 cloned into pFuse-IgG1 e3-Fc1 for the fragment containing the mutations. The successful exchange, resulting in PSG1 G93D95-S93L95, was confirmed by sequencing both strands of the cDNA.