关键词: 家蝇； 血细胞； MD14-3-3, 酵母双杂交； 互作蛋白

Abstract: 【Aim】To construct a yeast two-hybrid (Y2H) cDNA library of Musca domestica and investigate the proteins that could interact with the MD14-3-3. 【Methods】 Taking M. domestica as the research object, the mRNA was purified, the primary cDNA library was constructed, the secondary library was constructed by the recombinant pGADT7-DEST vector, so as to construct the Y2H. Then, the plasmid pGBKT7-MD14-3-3 were constructed as bait and used for screening the proteins interacting with MD14-3-3. Finally, the transformation and validation were verified. 【Results】 A Y2H cDNA library of M. domestica was successfully constructed, with the library storage capacity of 1.6×10⁷ CFU and the recombination rate of 100%. Two target proteins were obtained by Y2H screening. The results of transformation and validation showed that mucin-like protein HKR1 and antibacterial peptide ctenidin-1 interacted with MD14-3-3.【Conclusion】 Two proteins that interact with MD14-3-3 were identified from the successfully constructed Y2H cDNA library of M. domestica. Our results provide a basis for investigating the mechanism of immune response of M. domestica.

Key words: Musca domestica, hemocyte, MD14-3-3, yeast two-hybrid, interaction protein