The uncertainty of copy number variants: pregnancy decisions and clinical follow-up.

Background: Next-generation sequencing for copy number variants is often used as a follow-up investigation of unusual fetal ultrasound results and is capable of detecting copy number variations with a resolution of ∼0.1 Mb. In a prenatal setting, observation and subsequent management of pregnancies with a fetal variant of uncertain significance remains problematic for counseling.

Variant haplophasing by long-read sequencing: a new approach to preimplantation genetic testing workups.

Objective: To apply long-read, third-generation sequencing as a part of a general workup strategy for performing structural rearrangement (PGT-SR) and monogenic disease (PGT-M) embryo testing.

Design: Prospective study.

Setting: In vitro fertilization unit.

Third-generation sequencing: any future opportunities for PGT?

Purpose: To investigate use of the third-generation sequencing (TGS) Oxford Nanopore system as a new approach for preimplantation genetic testing (PGT).

Methods: Embryos with known structural variations underwent multiple displacement amplification to create fragments of DNA (average ~ 5 kb) suitable for sequencing on a nanopore.

Results: High-depth sequencing identified the deletion interval for the relatively large HBA1/2--SEA alpha thalassemia deletion.

Single-Molecule Sequencing: A New Approach for Preimplantation Testing and Noninvasive Prenatal Diagnosis Confirmation of Fetal Genotype.

We investigated the potential of next-generation sequencing (NGS) as an alternative method for preimplantation genetic testing of monogenic disease (PGT-M) with human leukocyte antigen (HLA) matching and for noninvasive prenatal diagnosis follow-up. The case involved parents who were carriers of the Fanconi anemia complementation group G (FANCG) 260delG mutation. After clinical PGT using conventional short tandem repeat and mutation analysis, two euploid disease-free embryos were transferred, resulting in a twin pregnancy.

Clinical application of single-molecule optical mapping to a multigeneration FSHD1 pedigree.

Introduction: Facioscapulohumeral muscular dystrophy 1 (FSHD1) is a relatively common autosomal dominant adult muscular dystrophy with variable disease penetrance. The disease is caused by shortening of a D4Z4 repeat array located near the telomere of chromosome 4 at 4q35. This causes activation of a dormant gene DUX4, permitting aberrant DUX4 expression which is toxic to muscles.

Preferential selection and transfer of euploid noncarrier embryos in preimplantation genetic diagnosis cycles for reciprocal translocations.

Objective: To develop and validate a new strategy to distinguish between balanced/euploid carrier and noncarrier embryos in preimplantation genetic diagnosis (PGD) cycles for reciprocal translocations and to successfully achieve a live birth after selective transfer of a noncarrier embryo.

Design: Retrospective and prospective study.

Setting: In vitro fertilization (IVF) units.

Prenatal diagnosis of chromosomal mosaicism in over 1600 cases using array comparative genomic hybridization as a first line test.

Objective: The aim of this study was to assess the detection of chromosomal mosaicism in chorionic villus (CVS) and amniotic fluid (AF) samples using array comparative genomic hybridization (aCGH) and quantitative fluorescent polymerase chain reaction.

Methods: All patients undergoing invasive prenatal testing by aCGH at a specialist prenatal screening service were included in the study. A total of 1609 samples (953 CVS and 656 AF) underwent quantitative fluorescent polymerase chain reaction and targeted aCGH without concurrent conventional G-banded karyotyping.

The genetic screening of preimplantation embryos by comparative genomic hybridisation.

Comparative genomic hybridization (CGH) is an indirect DNA-based test which allows for the accurate analysis of aneuploidy involving any of the 24 types of chromosomes present (22 autosomes and the X and Y sex chromosomes). Traditionally, embryos have been screened using fluorescence in situ hybridization (FISH)--a technique that was limited in the number of chromosomes able to be identified in any one sample. Early CGH reports on aneuploidy in preimplantation embryos showed that any of the 24 chromosomes could be involved and so FISH methods were going to be ineffective in screening out abnormal embryos.

What next for preimplantation genetic screening (PGS)? Experience with blastocyst biopsy and testing for aneuploidy.

Blastocysts more commonly have a normal karyotype than cleavage-stage embryos do. Moreover, blastocysts have also made a metabolic transition from catabolism and recycling of the oocyte's reserves and resources, processes that fuel the first 3 days of cleavage. Although not all blastocysts are karyotypically equal, it is still to be determined to what extent a mosaic karyotype might be a normal feature among embryos, both at the cleavage stage and the blastocyst stage--and when looking for karyotypic abnormalities by embryo biopsy might help the chance of implantation rather than harm it.

Pregnancies and live births after trophectoderm biopsy and preimplantation genetic testing of human blastocysts.

Objective: To compare multiple-cell trophectoderm biopsy for preimplantation genetic diagnosis (PGD) from day-5 blastocysts with previously published experience with day-3 cleavage-stage embryos.

Design: Retrospective review of laboratory and clinical experience.

Setting: Sydney IVF, a private clinic in Australia.

Moving to blastocyst biopsy for preimplantation genetic diagnosis and single embryo transfer at Sydney IVF.

Developments at Sydney IVF in the successful culture of blastocysts, combined with day 5 or 6 blastocyst biopsy and blastocyst cryostorage after biopsy, permit up to five or six cells to be genetically tested, leaving the inner cell mass intact and enabling embryos to be electively transferred one at a time. These advantages can be obtained without diminishing the chance of pregnancy compared with cleavage-stage biopsy and testing.