Insertional mutagenesis to isolate acetate-requiring mutants in Chlamydomonas reinhardtii

Abstract An arg 7 mutant of the green alga Chlamydomonas reinhardtii was transformed with pARG7.8, a plasmid bearing the wild-type ARG 7 gene. Out of 4100 arg⁺ transformants selected on an arginine-free medium supplemented with acetate, nine failed to grow on acetate-free medium (ac⁻ mutants). The results of the genetic and molecular analysis of several ac⁻ mutants are in agreement with the hypothesis that they originated from insertion of the incoming plasmid into the nuclear genome. These mutants should constitute valuable tools for isolating the corresponding wild-type genes after plasmid rescue into Escherichia coli .