Development and validation of a next-generation sequencing–based protocol for 24-chromosome aneuploidy screening of embryos

Fiorentino, Francesco; Biricik, Anıl; Bono, Sara; Spizzichino, Letizia; Cotroneo, Ettore; Cottone, Giuliano; Kokocinski, Felix; Michel, Claude-Edouard

doi:10.1016/j.fertnstert.2014.01.051

Cited by 223 publications

(185 citation statements)

References 33 publications

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“…While there are inherent differences in the genomic read length, the number of reads generated, and the algorithms used to precisely map the chromosomal position of sequencing reads, in principle, all methods generate a set of mapped reads for data analysis and allow direct comparisons to reference for identifying copy number changes. In validation studies against gold standard array comparative genomic hybridization [31,32], these methods have been proven to be very reliable and accurate for the detection of whole and partial chromosome aneuploidies. More recently, an NGS method based on a semiconductor ion torrent platform was developed and validated for PGD of translocations, with a reported resolution of 5 Mb for segmental imbalances [20].…”

Section: Discussionmentioning

confidence: 99%

“…This was achieved by incorporating two key modifications into the standard sequencing pipeline [25]. Firstly, we generated 2.8-3.2 million mapped sequencing reads of 36 bp in length, which is approximately threefold higher than that used by other published NGS methods developed for embryo chromosome analysis [20,31]. Secondly, sequencing reads are binned at 20-kb intervals (30-35 reads per bin), in contrast to 1-Mb chromosomal intervals used in the data analysis of other NGS methods.…”

Section: Discussionmentioning

confidence: 99%

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Clinical application of next-generation sequencing in preimplantation genetic diagnosis cycles for Robertsonian and reciprocal translocations

Zhang

Liu

Wang

et al. 2016

J Assist Reprod Genet

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Purpose The purpose of this study was to apply nextgeneration sequencing (NGS) technology to identify chromosomally normal embryos for transfer in preimplantation genetic diagnosis (PGD) cycles for translocations. Methods A total of 21 translocation couples with a history of infertility and repeated miscarriage presented at our PGD clinic for 24-chromosome embryo testing using copy number variation sequencing (CNV-Seq).Results Testing of 98 embryo samples identified 68 aneuploid (69.4 %) and 30 (30.6 %) euploid embryos. Among the aneuploid embryos, the most common abnormalities were segmental translocation imbalances, followed by whole autosomal trisomies and monosomies, segmental imbalances of nontranslocation chromosomes, and mosaicism. In all unbalanced embryos resulting from reciprocal translocations, CNV-Seq precisely identified both segmental imbalances, extending from the predicted breakpoints to the chromosome termini. From the 21 PGD cycles, eight patients had all abnormal embryos and 13 patients had at least one normal/balanced and euploid embryo available for transfer. In nine intrauterine transfer cycles, seven healthy babies have been born. In four of the seven children tested at 18 weeks gestation, the karyotypes matched with the original PGD results. Conclusion In clinical PGD translocation cycles, CNV-Seq displayed the hallmarks of a comprehensive diagnostic technology for high-resolution 24-chromosome testing of embryos, capable of identifying true euploid embryos for transfer.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Clinical application of next-generation sequencing in preimplantation genetic diagnosis cycles for Robertsonian and reciprocal translocations

Zhang

Liu

Wang

et al. 2016

J Assist Reprod Genet

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“…Next generation sequencing (NGS) is a technology that requires optimized DNA amplification to reduce the introduction of artifacts during the amplification process [49,50]. Following the DNA amplification, artifacts can be identified and removed by bioinformatics software.…”

Section: Next Generation Sequencingmentioning

confidence: 99%

Preimplantation genetic testing for aneuploidy: what technology should you use and what are the differences?

Brezina

Anchan

Kearns

2016

J Assist Reprod Genet

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Purpose The purpose of the review was to define the various diagnostic platforms currently available to perform preimplantation genetic testing for aneuploidy and describe in a clear and balanced manner the various strengths and weaknesses of these technologies. Methods A systematic literature review was conducted. We used the terms "preimplantation genetic testing," "preimplantation genetic diagnosis," "preimplantation genetic screening, " "preimplantation genetic diagnosis for aneuploidy," "PGD," "PGS," and "PGD-A" to search through PubMed, ScienceDirect, and Google Scholar from the year 2000 to April 2016. Bibliographies of articles were also searched for relevant studies. When possible, larger randomized controlled trials were used. However, for some emerging data, only data from meeting abstracts were available. Results PGS is emerging as one of the most valuable tools to enhance pregnancy success with assisted reproductive technologies. While all of the current diagnostic platforms currently available have various advantages and disadvantages, some platforms, such as next-generation sequencing (NGS), are capable of evaluating far more data points than has been previously possible. The emerging complexity of different technologies, especially with the utilization of more sophisticated tools such as NGS, requires an understanding by clinicians in order to request the best test for their patients.. Conclusion Ultimately, the choice of which diagnostic platform is utilized should be individualized to the needs of both the clinic and the patient. Such a decision must incorporate the risk tolerance of both the patient and provider, fiscal considerations, and other factors such as the ability to counsel patients on their testing results and how these may or may not impact clinical outcomes.

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“…This is because the same whole genome amplified product can be tested on different platforms (Fiorentino et al 2014) and the impact on the results of altered conditions of analysis (for example hybridization time on arrays) can be verified. The shift to trophectoderm biopsy at blastocyst stage has meant the effect of mosaicism in the sample needs to be validated for each new platform (Mamas et al 2012).…”

Section: Follow Up Of Untransferred Embryosmentioning

confidence: 99%

Quality control standards in PGD and PGS

SenGupta

Dhanjal

Harper

2016

Reproductive BioMedicine Online

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The aim of PGD is to test the preimplantation embryo for specific conditions for couples at risk of transmitting that genetic abnormality to their offspring.The couple need to go through IVF procedures to generate embryos in vitro.The embryos can be biopsied at either the zygote, cleavage or blastocyst stage. PGS uses the same technology to screen for chromosome abnormalities in embryos from patients going through IVF procedures as a method of embryo selection. Chromosome analysis was originally performed using fluorescent in situ hybridisation (FISH) which has now been replaced by array comparative genomic hybridisation or next generation sequencing (NGS) For the diagnosis of single gene defects, the polymerase chain reaction (PCR) has been used and has become highly developed over the years. More recently, SNP arrays and karyomapping have been introduced. PGD/PGS require partnership between IVF laboratories and diagnostic centres. As diagnosis can be performed using a variety of strategies with different technologies, accreditation of PGD diagnostic laboratories is important. Accreditation gives IVF centres an assurance that the diagnostic tests conform to specified standards. ISO 15189 is an international laboratory standard specific for medical laboratories. A requirement for accreditation is to participate in external quality assessment schemes Key wordsQuality control, Preimplantation genetic diagnosis 3 The current status of PGD and PGSInitial clinical application of PGD PGD was first performed in 1989 and since this time, genetic testing has seen major advances. PGD was developed as an alternative to prenatal diagnosis, for couples at risk of transmitting a genetic abnormality to their children.Couples have to go through IVF procedures to generate embryos in vitro, even though many of the couples that go through PGD are fertile. The embryos can be biopsied by the embryologists at the zygote stage (removal of the first and second polar body), cleavage stage (removal of 1-2 blastomeres from the 6-8 cell embryo) and blastocyst stage (removal of some trophectoderm cells) (Harton et al., 2011a). Up until recently, almost all PGD cycles were performed on blastomeres after cleavage stage biopsy (Harper et al, 2012, Moutou et al, 2014, but numerous studies have found that cleavage stage embryos exhibit high levels of chromosomal mosaicism, which means that biopsied cells may not be representative of the rest of the embryo (Harper et al, 1995, Munne et al, 1995, Fragouli et al, 2011, Taylor et al, 2014a. This is especially important when trying to perform PGD for a chromosome abnormality. Polar body biopsy is rarely used as it only gives genetic information on the maternal genome. In recent years, the IVF community have seen an increase in the use of blastocyst transfer (Glujovsky et al, 2012) and this has been reflected in the increased use of blastocyst biopsy for PGD (Moutou et al, 2014).The genetic testing should be performed by a specialised genetic testing laboratory. The very first cases of PGD wer...

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Development and validation of a next-generation sequencing–based protocol for 24-chromosome aneuploidy screening of embryos

Cited by 223 publications

References 33 publications

Clinical application of next-generation sequencing in preimplantation genetic diagnosis cycles for Robertsonian and reciprocal translocations

Clinical application of next-generation sequencing in preimplantation genetic diagnosis cycles for Robertsonian and reciprocal translocations

Preimplantation genetic testing for aneuploidy: what technology should you use and what are the differences?

Quality control standards in PGD and PGS

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