A FORWARD GENETIC SCREEN FOR ISOLATION OF FUSION DEFECTIVE MATING TYPE MINUS MUTANTS IN CHLAMYDOMONAS REINHARDTII

Abstract

Many biological processes require cell fusion, therefore defects in cell fusion result in many diseases. The unicellular green alga Chlamydomonas reinhardtii is an excellent model organism to study cell-cell fusion. The objective of this study was to identify genes that are involved in C. reinhardtii mating type minus (MT_) gamete fusion. A forward genetics approach was taken in our work. We created several MT_ fusion defective mutants using DNA insertional mutagenesis. These mutants were normal in the early stages of mating; they agglutinated with mating type plus (MT+), removed their walls, adhered to their mating partner through their mating structures, but the cells did not fuse, indicating that the DNA insertional mutants were defective in the latest stages of fusion. The number of insertions was confirmed by Southern blots. Mutant J1 had one insertion and the flanking genomic DNA was cloned by TAIL-PCR and RESDA-PCR. The insertion is in a gene predicted to be involved in 5-deoxystrigol biosynthesis.