Study On The Major Royal Jelly Protein Genes From The Chinese Honeybee And Their Expression In E. Coli And Insect Cells

The honeybee, a model organism for studying insect social behavior, is well-studied physiologically. It has been reported that Royal Jelly (RJ) has several pharmacological activities. Nowadays, scientists begin to pay attention to the molecular characterization of RJ, which plays a major role in honeybee queen development as larvae are non selectively fed for the first three days, but only the larvae that developed into queen bees are continually fed RJ throughout their lives. So RJ plays a major role in caste differentiation.A substantial part of RJ is proteins, which form about 46%-89% of the content of RJ. MRJPs (Major Royal Jelly Proteins) were identified as major protein components of royal jelly, constituting about 90% of the total protein content. It is found that MRJPs belong to one protein family (60%-70% amino acid sequence homology between the proteins). Biological activities of Apis mellifera MRJPs, such as AmMRJP1 and AmMRJP3, have been studied in various systems. Rearing experiments have illustrated that Apis mellifera queens were not successfully reared with Apis cerara RJ, and vice versa. This implied possible differerces of the potential compositions of the RJ from Apis mellifera and Apis cerana. But compare to Apis mellifera, information on MRJPs of Apis cerana is scarce.We have constructed a cDNA library from nurse honeybee heads of Apis cerana cerana, and the clones of AccMRJP7, AccMRJP5 and AccMRJP3a genes were screened out. The target gene fragment of AccMRJP7 was expressed in E. coli BL21(DE3) successfully, but the products existed mainly as inclusion body. We purified the protein and finished the antibody preparation of AccMRJP7. The titer and specificity of the antibody was analyzed by ELISA and Western blotting. The expressed AccMRJP7 reached to 22.8% of the total proteins. AccMRJP5 and AccMRJP3a genes were expressed in Eukaryotic expression system. Western blot analysis confirmed that AccMRJP3a and AccMRJP5 genes were successfully expressed in the Bac to Bac system. The product of AccMRJP5 gene only displayed one specifically expressed band, but AccMRJP3a gene displayed three. The expressed AccMRJP5 and AccMRJP3a reached to 15.1% and 14.51% of the total proteins, respectively.