Multiple genotyping in bovine pre‐implantation embryos with whole genome amplification

This study examined genetic diagnosis using whole genome amplification (WGA) in bovine embryos. The first experiment was conducted to compare the WGA efficiency of primer extension preamplification-PCR (PEP-PCR) and multiple displacement amplification (MDA), and to optimize the DNA extraction method. The sensitivity of SRY-specific PCR from MDA products increased when DNA of fibroblasts was extracted by a NaOH treatment instead of the conventional method (heat treatment). The detectability of SRY from PEP-PCR products was lower than that in MDA regardless of the DNA extraction method (proteinase K or NaOH treatment). Sexing and genotyping were performed using MDA products from embryo biopsy. The accuracy of PCR-based and LAMP-based sexing was 100% regardless of the amounts of biopsy. Genotyping of CL16, BND3, SCD and F11 in MDA products from 10 to 20% of trophectoderm was successful 97, 97, 95 and 95% of the time, respectively, but reduced biopsy amount (<10% of trophectoderm) decreased the accuracies (33–83%). Microsatellite markers were analyzed using MDA products from 10 to 20% of trophectoderm. In eight out of 16 microsatellites, genotypes were not contradictory among the dam, sire and embryos. In the other eight microsatellites, the inconsistency rates were 17–83%. These results indicate that MDA is useful for multiple genetic diagnoses in bovine embryos.