Molecular genetic characterization of new bovine kappa-casein alleles CSN3(F) and CSN3(G) and genotyping by PCR-RFLP

In order to characterize the two new kappacasein variants F and G (CSN3(F) and CSN3(G)) recently detected in Ayrshire and Pinzgauer cattle, exon IV of CSN3 from heterozygous animals was amplified by polymerase chain reaction (PCR), cloned and sequenced. The sequencing data revealed single point mutations at nucleotide positions 10530 (G-->A) for CSN3(F) and 10790 (C-->T) for CSN3(G), corresponding to amino acid exchanges in positions 10 (Arg-->His) and 97 (Arg-->Cys) respectively. These mutations alter recognition sites for the restriction enzymes HhaI and MaeII, which were subsequently used to confirm these polymorphisms in cattle carrying CSN3(F) or CSN3(G). A PCR-restriction fragment length polymorphism (RFLP) genotyping procedure for all currently known CSN3 alleles (CSN3(A), CSN3(B), CSN3(C), CSN3(E), CSN3(F), CSN3(G)) was developed.