Laboratory Culture and Mutagenesis of Amphioxus (Branchiostoma floridae).

Cephalochordates (amphioxus) are invertebrate chordates closely related to vertebrates. As they are evolving very slowly, they are proving to be very appropriate for developmental genetics studies aimed at understanding how vertebrates evolved from their invertebrate ancestors. To date, techniques for gene knockdown and overexpression have been developed, but methods for continuous breeding cultures and generating germline mutants have been developed only recently. Here we describe methods for continuous laboratory breeding cultures of the cephalochordate Branchiostoma floridae and the TALEN and Tol2 methods for mutagenesis. Included are strategies for analyzing the mutants and raising successive generations to obtain homozygotes. These methods should be applicable to any warm water species of cephalochordates with a relatively short generation time of 3-4 months and a life span of 3 years or more.