Technical note: Development and application of KASP assays for rapid screening of 8 genetic defects in Holstein cattle

Abstract

Specific DNA mutations underlying several genetic defects associated with embryo loss or reduced calf survivability have been identified in dairy cattle, and a convenient and cost-effective platform is required for their routine screening. We developed Kompetitive allele-specific PCR (KASP) assays for discrimination of the wild-type alleles from the associated defective alleles at each of 8 common genetic defects in Holstein cattle, involving 5 SNP [HH1, HH3, HH4, bovine leukocyte adhesion deficiency (BLAD), and complex vertebral malformation (CVM)] and 3 insertion or deletion mutations [HH5, haplotype for cholesterol deficiency (HCD), and brachyspina (BS)]. A total of 390 cows from a Chinese Holstein herd were genotyped and the carriers identified at 7 of these 8 loci (except HH4), with the highest carrier frequencies found for CVM (10.5%) and HH1 (10.0%), followed by HH3 (2.6%), BS (2.1%), HCD (1.3%), HH5 (0.8%), and BLAD (0.5%). Surprisingly, 102 cows (26.2%) carried at least 1 of the 7 defective alleles. Our results demonstrate that these KASP assays are simple, rapid, and reliable for the detection of multiple genetic defects. The high carrier frequency of these genetic defects indicates an urgent need for routine molecular testing to eliminate the deleterious alleles from Chinese Holstein cattle.

Genotype Screening of Bovine Brachyspina in Taiwan Holstein Cows

“…BY carriers were found in 17 of 31 herds, with 1-25 cows in each herd, with carrier frequencies of 1.85-15%. The total carrier frequency of 5.09% was lower than that reported in the United States (6%) (VanRaden et al, 2011), the Netherlands (7.4%) (Charlier et al, 2012) and Poland (10.26%) (Ruść and Kaminski, 2015) and higher than that reported in China (2.6%) (Zhang et al, 2020). It was also lower than that reported by previous Taiwanese studies, in which 409, 172 and 436 cattle from 4, 4 and 9 herds, respectively, with BY carrier frequencies of 8.56, 12.21 and 6.65%, respectively, were sampled (Liaw et al, 2015;2017;Lai et al, 2017).…”

“…Effective screening to prevent further transmission of the BS allele is necessary. For simultaneous genetic screening of defective alleles in Holstein cows, Zhang et al (2020) developed the Kompetitive Allele-Specific Polymerase chain reaction (KASP) assay for discrimination of wild-type alleles from eight defective alleles associated with Holstein cows. However, the KASP assay is only appropriate for analysis of small to moderate numbers of loci.…”

“…Multiplexed, chip-based, high-throughput genotyping technology is preferred for analysis of large numbers of genomic Single-Nucleotide Polymorphisms (SNPs) (Zhang et al, 2020). The Igenity-Prime 50 k SNP chip, which comprises nearly 42,000 genetic markers, is cost-effective, provides a comprehensive genomic profile and yields highly accurate results.…”

Prevalence of nine genetic defects in Chinese Holstein cattle

“…Notably, the recently developed microfluidic chip combining with KASP assay has made it possible to screen different loci simultaneously (Lu et al., 2019). This microfluidic method has been applied for molecular screening of eight common genetic defects (BLAD, BS, CVM, HH1, HH3, HH4, HH5 and HCD) in Holstein cattle (Zhang et al., 2020).…”

“…This study aimed to develop a KASP assay for genotyping the HH6 locus, to integrate this assay with the method already developed for the eight genetic defects (BLAD, BS, CVM, HH1, HH3, HH4, HH5 and HCD) that are common in Holstein cattle (Zhang et al., 2020) and to determine the frequencies of carriers of each of these nine defects in Chinese Holstein using more samples than the previous study.…”