Practical use of DNA polymorphisms in the avian immunoglobulin light chain constant domain for species-specific PCR

摘要

The species-specific polymorphisms in the immunoglobulin light chain (IgL) gene of chickens, pheasants (Phasianus colchicus), quail (Coturnix japonica), and turkey (Meleagris gallopavo) were examined. Species specific primers were designed for species-specific PCR. Additional species-specific primers were designed for cytochrome b and tapasin genes for purposes of comparison with IgL primers. Primers specific for the chicken IgLV were successfully used to amplify DNA from chicken (control), pheasant, turkey, and quails. The amplified products were cloned and sequenced. The sequences were very similar in framework regions 2 and 3. Cross amplification of the IgLV between species using the same primers indicated that both ends of the IgLV were similar among the members of the Phasianidae family and that the primers could be used to clone the IgLV in other avian species. The alignment of the IgLC from each species indicated a high level of conservation, the level of homology between each pair of species was 83-95%. The potential for species identification based on inter-species polymorphisms in the IgLC was confirmed by PCR using species-specific primer sets which annealed to pheasant, turkey or quail, but not chicken DNA due to a sequence difference in the IgLC. The primer set PSP-tapasin was designed based on the sequence difference between pheasant and chicken involving a 95 pb insertion in pheasant tapasin, and the primer set TSP-tapasin which was specific for turkey was designed on the 8 bp deletion in turkey tapasin. It is concluded that species-specific primer sets are species-specific and can be used to screen for interspecies germline chimaeras between chicken and other avian species.