The generation of antibody diversity in the turtle.
The Ab response in reptiles has been studied at the protein level, and in turtles some aspects resemble those of cold-blooded vertebrates from other classes. The genetic bases for these features are not clear. The present study is the first on the IgH organization and complexity of a reptilian Ig gene system. The approach to cloning turtle (Pseudemys scripta) sequences is entirely PCR based, and its efficacy is demonstrated by obtaining extensive information on a heretofore unexplored Ig gene system. A number of genomic VH sequences, representing possibly four families, were isolated, as was a genomic C mu 4 clone. These sequences, used as probes, provided proof that in the turtle there is a single IgH locus with multiple VH genes and one C mu gene. In Northern hybridizations, the C mu 4 probe detected two transcripts; of the four VH groups, only one was expressed, and multiple bands indicated the presence of at least two non-mu transcripts. Using reverse transcription-PCR on spleen or liver RNA, an IgM heavy chain sequence was obtained, as were a number of VDJ rearrangements. Among 32 unique VDJ rearrangements from one animal, there were 22 sequence variants at framework 4, suggesting either a very large number of J segments or somatic modification in the variable region. The latter interpretation is supported by point mutations found in framework 3 and CDR3. The number of changes is considerably greater than the deduced Taq misincorporation rate (0.05%).
Turchin
A
A
Department of Biology, New York University 10003, USA.
Hsu
E
E
D000595:Amino Acid Sequence; D000818:Animals; D000916:Antibody Diversity / Q000235:genetics*; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D005803:Genes, Immunoglobulin; D007143:Immunoglobulin Heavy Chains / Q000235:genetics / Q000302:isolation & purification; D007135:Immunoglobulin Variable Region / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D005810:Multigene Family; D016133:Polymerase Chain Reaction; D014426:Turtles
D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region