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Biomed Res Int. 2017;2017:1209158. doi: 10.1155/2017/1209158. Epub 2017 Jun 22.

Whole Genome Amplification of Day 3 or Day 5 Human Embryos Biopsies Provides a Suitable DNA Template for PCR-Based Techniques for Genotyping, a Complement of Preimplantation Genetic Testing.

Author information

1
Laboratorio de Investigación y Diagnóstico Molecular, Instituto de Infertilidad y Genética, Ingenes, Ciudad de México, Mexico.
2
Departamento de Genética y Biología Molecular, Cinvestav-IPN, Ciudad de México, Mexico.
3
Departamento de Toxicología, Cinvestav-IPN, Ciudad de México, Mexico.

Abstract

Our objective was to determine if whole genome amplification (WGA) provides suitable DNA for qPCR-based genotyping for human embryos. Single blastomeres (Day 3) or trophoblastic cells (Day 5) were isolated from 342 embryos for WGA. Comparative Genomic Hybridization determined embryo sex as well as Trisomy 18 or Trisomy 21. To determine the embryo's sex, qPCR melting curve analysis for SRY and DYS14 was used. Logistic regression indicated a 4.4%, 57.1%, or 98.8% probability of a male embryo when neither gene, SRY only, or both genes were detected, respectively (accuracy = 94.1%, kappa = 0.882, and p < 0.001). Fluorescent Capillary Electrophoresis for the amelogenin genes (AMEL) was also used to determine sex. AMELY peak's height was higher and this peak's presence was highly predictive of male embryos (AUC = 0.93, accuracy = 81.7%, kappa = 0.974, and p < 0.001). Trisomy 18 and Trisomy 21 were determined using the threshold cycle difference for RPL17 and TTC3, respectively, which were significantly lower in the corresponding embryos. The Ct difference for TTC3 specifically determined Trisomy 21 (AUC = 0.89) and RPL17 for Trisomy 18 (AUC = 0.94). Here, WGA provides adequate DNA for PCR-based techniques for preimplantation genotyping.

PMID:
28717645
PMCID:
PMC5498896
DOI:
10.1155/2017/1209158
[Indexed for MEDLINE]
Free PMC Article

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