Structure and genomic organization of VH gene segments in the channel catfish: Members of different VH gene families are interspersed and closely linked

Abstract

To determine the structure and organization of germline VH gene segments in the channel catfish, genomic lambda libraries were screened with cDNA probes representing different catfish VH gene families. Thirty-six VH positive genomic clones were isolated and four of these were characterized by restriction mapping and Southern blot analysis with probes specific for each known VH gene family. The four clones, representing about 65 kb of DNA, contained 21 VH segments. The average distance between segments was about 3 kb and gene segments representing different VH gene families were interspersed with each other. Dot-blot hybridization analysis of all 36 genomic clones (average insert size 16–18 kb) indicated that the average clone contained gene segments representing four different VH families. In addition, these analyses indicated that VH segments representing each VH family could be found closely linked to gene segments representing each of the other VH families. Genomic restriction fragments containing a VH segment of each gene family were sequenced. These analyses showed that the general structure of VH segments is conserved in catfish. These structural features include the presence of a leader sequence split by a short intron, an uninterrupted open reading frame encoding readily identified framework and complementarity determining regions, and a downstream recombination signal sequence represented by a consensus heptamer, a22–24 bp spacer, and an A-rich nonamer. Upstream of the VH segments was an octamer sequence. These analyses indicate that the organization and structure of VH segments typically associated with VH loci of higher vertebrates evolved early in phylogeny at the level of the bony fishes.