Student Research Project
Study of reptilian immunoglobulin gene system

Student
Aleksandr Turchin
Major: Biology
Future Plans: Medical school

Professor
Ellen Hsu, Assistant Professor, Department of Biology, New York University, New York

This research project investigates the basic principles of organization of the immunoglobulin gene system in reptiles. Immunoglobulins (antibodies) are protein molecules that are synthesized by cells of the immune system (B lymphocytes). Their function is to recognize substances foreign to the organism and help eliminate them. "Variable" and "constant" regions can be distinguished in the antibody molecule. The variable region recognizes the substance against which this particular immunoglobulin is directed (there are many such substances, so there are many versions of the variable region, hence the name), and the constant region has an effector function, which plays the role of a marker indicating to the cells that eliminate potential pathogens that the substance to which the antibody is bound has to be destroyed.

An immunoglobulin molecule is encoded by several genes, which can be organized in various ways, resulting in different mechanisms of assembly of the whole immunoglobulin molecule. A diversity of specificities must be generated to cover a wide range of foreign substances. So far, immunoglobulin genes have been investigated in all vertebrate classes except reptiles. The goal of our project is to clone immunoglobulin genes from a reptile and analyze the gene structure and the mechanisms of diversity generation.

We use turtle genomic DNA as a source of genetic material from which to clone the gene. Since the concentration of the gene of interest in the genome is very small, we employ polymerase chain reaction to amplify it. Afterwards, the gene is inserted into a plasmid and sequenced. The sequence is then compared to the known sequences of immunoglobulin genes from other species. To test for the presence of the cloned sequence in the turtle genome and to determine the number of homologous genes, Southern blotting with turtle genomic DNA is performed using the cloned sequence as a probe.

We have isolated a number of sequences we identify as encoding portions of variable and constant regions of the immunoglobulin molecule on the basis of the presence of key elements of immunoglobulin gene structure and derived amino acid sequence. We have also demonstrated the presence of multiple genes encoding heavy chain variable region and no more than one gene encoding heavy chain constant region in the turtle genome. These findings indicate that the reptilian immunoglobulin gene system resembles that of mammals as opposed, for example, to sharks.

The next step is to analyze the sequences of the "complementarity-determining regions," which would shed further light on the mechanism of antibody diversity generation in reptiles. In the future, we also intend to isolate individual genes comprising the immunoglobulin gene system from a genomic DNA library, which would allow us to study untranslated portions of the genes that have important regulatory function.

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