Genotyping Test Development and Genotyping Survey of Pakistani population of Holstein Friesian imported from Different Origins for A1/A2 SNP in Beta-casein Gene

Quality of milk is determined by multiple factors including microbial load, somatic cell score, content percentages and absence of adulterants. However, in term of genetics, milk quality can be linked with beta casein. Beta casein is a milk protein having genetic variants, one of which is believed to be linked with progression of many diseases including cardiovascular diseases, metabolic diseases and nervous disorders. As beta casein has genetic basis so milk quality can be elucidated by animal genotyping. But no such genotyping facility was available country wide for this purpose. Hence objective of this study is the development of diagnostic test using ARMS-PCR technology in Pakistan and genotyping of beta casein variants in Pakistani population of Holstein Friesian cattle. A total of 166 blood samples were collected from Holstein Friesian (imported from Holland, USA and Australia) and genotyped using ARMS-PCR. Validation of ARMS-PCR was confirmed by Sanger sequencing of beta casein gene from 78 animals. ARMS-PCR based test was then compared with parallel genotyping techniques in term of cost and time effectivity. Reproducibility of this test was tested and validation of this test was done by Sanger sequencing. This test is cost effective as well as time effective in compared to other techniques. Holstein Friesian originating from different countries have different percentages of beta casein variants. This test can be used routinely for genotyping of beta casein. In future there is a need to find out prevalence of beta casein variants of indigenous cattle as well as cross bred population of Pakistan and formulation of breeding policy, keeping in view health concerns of A1 genotype of {beta}-casein.\n\nNovelty StatementIn this study, a diagnostic test is developed to genotype beta casein variants in animal. This is the first report of its kind in Pakistan as before this report either sequencing or tedious and costly procedures were in use for genotyping of beta casein variants in Pakistan. Additionally, for the first time in Pakistan Holstein Friesian imported from Holland, United States and Australia were genotyped to elucidate their genotype status for beta casein polymorphism.