Abstract:To investigate the function of ARL3 in ciliogenesis and ciliary signal transduction, the author generate a rabbit polyclonal antibody against Chlamydomonas reinhardtii ARL3 and to identify arl3 mutants. The target gene, arl3, was obtained through gene cloning, and a prokaryotic expression vector, pET-28a(+)-arl3, was constructed for expression in Escherichia coli BL21 (DE3) to produce the fusion protein 6×His-ARL3. The purified target protein was used to immunize rabbits, resulting in the production of antiserum. The antiserum was subsequently purified using protein A and antigen-antibody, and Western blot was employed to detect the presence of the Chlamydomonas reinhardtii ARL3 antigen. Results indicated that the antiserum titer reached 1∶102 400, with the purified ARL3 antibody detecting a single specific band at a dilution of 1∶400. This confirms the successful preparation of a highly specific and sensitive rabbit polyclonal antibody against Chlamydomonas reinhardtii ARL3, as well as the identification of an arl3 mutant. These findings provide a foundational basis for further investigation into the function of ARL3 protein within cilia.
