Cloning And Immunologic Recognition Of Antigen Desmoglein 4 Fragments In Pemphigus Vulgaris

Pemphigus is an autoimmune disease that results in blistering of cutaneous and mucous membrane. It is caused by autoantibodies directed against desmoglein on keratinocytes, and pemphigus vulgaris (PV) is one of the subtypes which can be frequently seen. Although it has been proved that desmoglein l(Dsgl), desmoglein 3(Dsg3) and their autoantibodies are relevant to pemphigus onset, the precise pathogenesis is still dimness. Recent study showed that desmoglein 4 from human body is also the target antigen of autoantibodies in PV which provided a new clue to research the disease.The aim of the study is to amplify the nucleotide sequences of desmoglein 4(Dsg4) extracellular domains (ECs), as EC1, EC2, EC3 and EC4 (EC1-4) from human skin tissue, and then investigate their role in PV pathogenisis.Firstly, specificiticity primers were designed according to cDNA sequences of Dsg4 in Genebank data. RNA was obtained from normal human skin tissue and then cDNA was synthesized. Objective genes of desmoglein 4 extracellular domains (EC1-4) were amplified by polymerase chain reaction (PCR). With the technique of gene recombination, objective genes linkaged with pET32a plasmids respectively by T4DNA ligase, which translated into Escherichia coli (E.coli) DH5α competent germs. Screening of translated germs was done by Luria-bertani (LB) medium with Ampicillin. DNA sequences of positive recons were identified finally. The results showed that four bands with all the length of 350bp were obtained by RT-PCR. Consequently four expressed plasmids of desmoglein 4 extracellular domains...