Sequence analysis of members of the human Ig V_H4 gene family derived from a single V_H locus: Identification of novel germ-line members

We have generated a mouse × human heterohybridoma that contains a single copy of chromosome 14 and, thus, a haploid set of Ig V_H genes. This cell line was used to investigate the germ-line content and nucleotide sequences of members of the V_H4 gene family in a polymerase chain reaction-based approach. The analysis of 58 full-length sequences revealed the presence of 12 different germ-line V_H4 genes, each of which is potentially functional. These germ-line V_H4 genes were compared with the nucleotide sequences of published V_H4 genes. Three V_H4 genes were 100% identical to previously published sequences and belong to a group of V_H4 genes that are strongly conserved and highly prevalent in the human population. Three V_H4 genes in our collection displayed >99.3% sequence identity with reported germ-line V_H4 sequences and likely represent allelic counterparts of these genes. Six genes displayed <97.2% sequence identity with published V_H4 genes and were identified as novel members of the human V_H4 gene family or more distantly related alleles of known V_H4 genes. Collectively, these data suggest that, overall, the human V_H4 gene family may be more diverse than hitherto assumed, whereas a number of individual members are non-polymorphic and extremely well conserved.