PROKARYOTIC EXPRESSION, PURIFICATION AND POLYCLONAL ANTIBODY PREPARATION OF CHLAMYDOMONAS REINHARDTII BBS8

The purpose of this study is to prepare a highly specific rabbit polyclonal antibody against Chlamydomonas reinhardtii bbs8, and lay a material foundation for the follow-up study of BBS8 protein function. To achieve this, N-terminal 6×His-tagged BBS8 fusion protein (6×His::BBS8) was expressed in E. coli and purified with Ni-column. New Zealand white rabbits were immunized three times with 6×His::BBS8. Antiserum were used for ELISA at 1:102400 dilution. The IgG subtypes were enriched with protein A resin, which were affinity-purified with the purified E. coli-expressed N-terminal MBP-tagged BBS8 (MBP::BBS8) protein. The anti-BBS8 were validated using whole cell extracts of C. reinhardtii CC-125 and BBS8-null mutant bbs8 strains with high specificity, thus it is suitable for investigating the function of BBS8.