摘要
In order to study the DLA(Dog Leucocyte Antigen)classⅡ region we utilized the polymerase chain reaction based restriction fragment length polymorphism(FCR-RFLF)method,whick has been reported previously as an efficient and simple technique for accurate definition of the HLA class Ⅱ alleles[1].To search for a appropriate primer pair a serie of amplifications with 4 different primer pairs DLA-DR-SP-Stop,DLA-DR-SP/P3,HLA-DRB-GH46/50 and HLA-DRB-AMF-A/B were provided.Only one satisfactory amplification was obtained with the primer pair HLA-DRB-AMP-A/E.The anelogous sequences of the primer pair are found in the sequence of HLA-DRB-cDNA.The amplification region of the primer pair includets also the three hypervariable regions(HVR)in the seqtence of DLA-DRB cDNA. Southern blot hybridization analysis confirmed the specificity of the primer pair HLA-DRB-AMP-A/B.The results of Hae Ⅲ and Hinfl digestion show high polymorphism in DLA-D region and allele specific polymorphic patterns.Therefore,it is suggested that the primer pair HLA-DRBAMP-A/E is at present the only available and usefull primer pair in FCR-RFLP stuty of DLA-DRB1 gene.
In order to study the DLA(Dog Leucocyte Antigen)classⅡ region we utilized the polymerase chain reaction based restriction fragment length polymorphism(FCR-RFLF)method,whick has been reported previously as an efficient and simple technique for accurate definition of the HLA class Ⅱ alleles[1].To search for a appropriate primer pair a serie of amplifications with 4 different primer pairs DLA-DR-SP-Stop,DLA-DR-SP/P3,HLA-DRB-GH46/50 and HLA-DRB-AMF-A/B were provided.Only one satisfactory amplification was obtained with the primer pair HLA-DRB-AMP-A/E.The anelogous sequences of the primer pair are found in the sequence of HLA-DRB-cDNA.The amplification region of the primer pair includets also the three hypervariable regions(HVR)in the seqtence of DLA-DRB cDNA. Southern blot hybridization analysis confirmed the specificity of the primer pair HLA-DRB-AMP-A/B.The results of Hae Ⅲ and Hinfl digestion show high polymorphism in DLA-D region and allele specific polymorphic patterns.Therefore,it is suggested that the primer pair HLA-DRBAMP-A/E is at present the only available and usefull primer pair in FCR-RFLP stuty of DLA-DRB1 gene.
