title: Accuracy of preimplantation genetic diagnosis in equine in vivo-recovered and in vitro-produced blastocysts.
abstract: Preimplantation genetic diagnosis has great potential in the horse, but information on evaluation of equine embryo biopsy samples is limited. Blastocysts were biopsied using a Piezo drill and methods for whole-genome amplification (WGA) investigated. Results for 33 genetic loci were then compared between biopsy samples from in vitro-produced (IVP) and in vivo-recovered (VIV) blastocysts. Under the experimental conditions described, WGA using the Qiagen Repli-g Midi kit was more accurate than that using the Illustra Genomiphi V2 kit (98.2% vs 25.8%, respectively). Using WGA with the Qiagen kit, three biopsy samples were evaluated from each of eight IVP and 19 VIV blastocysts, some produced using semen from stallions carrying the genetic mutations associated with the diseases hereditary equine regional dermal asthenia (HERDA), hyperkalemic periodic paralysis (HYPP) or polysaccharide storage myopathy 1 (PSSM1). Three of 81 biopsy samples (3.7%) returned <50% accuracy. In the remaining 78 samples, overall accuracy was 99.3% (2556/2574 loci interrogated). Accuracy did not differ significantly between samples from IVP and VIV blastocysts. Allele drop-out in heterozygous loci was 1.6% (17/1035). Accuracy for sex determination was 100%; accuracy for heterozygosity for disease-causing mutations was 97.7% (43/44). In conclusion, Piezo-driven embryo biopsy with WGA has >95% overall accuracy in IVP and VIV embryos, and this technique is suitable for use in a clinical setting.
authors.affiliation: Department of Veterinary Physiology and Pharmacology, Texas A&M University, 4466 TAMU, College Station, TX 77843-4466, USA.
authors.forename: Y H
authors.initials: YH
authors.lastname: Choi
authors.affiliation: Veterinary Genetics Laboratory, University of California, Old Davis Road, Davis, CA 95616, USA.
authors.forename: M C T
authors.initials: MCT
authors.lastname: Penedo
authors.affiliation: Veterinary Genetics Laboratory, University of California, Old Davis Road, Davis, CA 95616, USA.
authors.forename: P
authors.initials: P
authors.lastname: Daftari
authors.affiliation: Department of Veterinary Physiology and Pharmacology, Texas A&M University, 4466 TAMU, College Station, TX 77843-4466, USA.
authors.forename: I C
authors.initials: IC
authors.lastname: Velez
authors.affiliation: Department of Veterinary Physiology and Pharmacology, Texas A&M University, 4466 TAMU, College Station, TX 77843-4466, USA.|Department of Large Animal Clinical Sciences, Texas A&M University, 4475 TAMU, College Station, TX, 77843-4475, USA.|Corresponding author. Email: khinrichs@cvm.tamu.edu.
authors.forename: K
authors.initials: K
authors.lastname: Hinrichs
keywords: HERDA; HYPP; PSSM; embryo biopsy; genetic screening; polymerase chain reaction (PCR); whole genome amplification