A Decade of Molecular Preimplantation Genetic Diagnosis of 350 Blastomeres for Beta-Thalassemia Combined with HLA Typing, Aneuploidy Screening and Sex Selection in Iran

Abstract Background: Preimplantation genetic diagnosis (PGD) has been developed to detect genetic disorders before pregnancy which is usually done on blastomeres biopsied from 8-cell stage embryos obtained from in vitro fertilization method (IVF).Here we report molecular PGD results for diagnosing of beta thalassemia (beta-thal) which are usually accompanied with evaluating chromosomal aneuploidies, HLA typing and sex selection.Methods: In this study, haplotype analysis was performed using short tandem repeats (STRs) in a multiplex nested PCR and the causative mutation was detected by Sanger sequencing.Results: We have performed PGDs on 350 blastomeres from 55 carrier couples; 142 blastomeres for beta-thal only, 75 for beta-thal and HLA typing, 76 for beta-thal in combination with sex selection, and 57 for beta-thal and aneuploidy screening. 150 blastomeres were transferable, 15 pregnancies were happened, and 11 babies born.We used 6 markers for beta-thal, 36 for aneuploidy screening, 32 for sex selection, and 35 for HLA typing. To our knowledge combining all these markers together and the number of STR markers are much more than any other studies which have ever done.Conclusions: PGD is a powerful diagnostic tool for carrier couples who desire to have a healthy child and wish to avoid medical abortion.

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Preimplantation Genetic Diagnosis—An Overview

Journal of Histochemistry & Cytochemistry ◽

10.1369/jhc.4b6395.2005 ◽

2005 ◽

Vol 53 (3) ◽

pp. 255-260 ◽

Cited By ~ 56

Author(s):

Caroline Mackie Ogilvie ◽

Peter R. Braude ◽

Paul N. Scriven

Keyword(s):

Preimplantation Genetic Diagnosis ◽

Genetic Diagnosis ◽

Sex Selection ◽

Hla Typing ◽

Success Rates ◽

Aneuploidy Screening ◽

Reciprocal Translocations ◽

Public Controversy ◽

Polar Bodies ◽

Monogenic Diseases

Since the early 1990s, preimplantation genetic diagnosis (PGD) has been expanding in scope and applications. Selection of female embryos to avoid X-linked disease was carried out first by polymerase chain reaction, then by fluorescence in situ hybridization (FISH), and an ever-increasing number of tests for monogenic diseases have been developed. Couples with chromosome rearrangements such as Robertsonian and reciprocal translocations form a large referral group for most PGD centers and present a special challenge, due to the large number of genetically unbalanced embryos generated by meiotic segregation. Early protocols used blastomeres biopsied from cleavage-stage embryos; testing of first and second polar bodies is now a routine alternative, and blastocyst biopsy can also be used. More recently, the technology has been harnessed to provide PGD-AS, or aneuploidy screening. FISH probes specific for chromosomes commonly found to be aneuploid in early pregnancy loss are used to test blastomeres for aneuploidy, with the aim of replacing euploid embryos and increasing pregnancy rates in groups of women who have poor IVF success rates. More recent application of PGD to areas such as HLA typing and social sex selection have stoked public controversy and concern, while provoking interesting ethical debates and keeping PGD firmly in the public eye.

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First successful pregnancy outcome after preimplantation genetic diagnosis for aneuploidy screening in embryos generated from natural-cycle in vitro fertilization combined with an in vitro maturation procedure

Fertility and Sterility ◽

10.1016/j.fertnstert.2005.10.066 ◽

2006 ◽

Vol 85 (5) ◽

pp. 1510.e9-1510.e11 ◽

Cited By ~ 13

Author(s):

Asangla Ao ◽

Shaoguang Jin ◽

Durga Rao ◽

Weon-Young Son ◽

Ri-Cheng Chian ◽

...

Keyword(s):

In Vitro Fertilization ◽

Pregnancy Outcome ◽

Preimplantation Genetic Diagnosis ◽

In Vitro Maturation ◽

Genetic Diagnosis ◽

Natural Cycle ◽

Successful Pregnancy ◽

Aneuploidy Screening ◽

Vitro Fertilization

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A versatile strategy for preimplantation genetic diagnosis of haemophilia A based on F8-gene sequencing

Thrombosis and Haemostasis ◽

10.1160/th06-05-069 ◽

2006 ◽

Vol 96 (12) ◽

pp. 839-845 ◽

Cited By ~ 10

Author(s):

Jorge Sánchez-García ◽

Dominique Gallardo ◽

Joaquima Navarro ◽

Carmen Márquez ◽

Josep Gris ◽

...

Keyword(s):

Preimplantation Genetic Diagnosis ◽

Clinical Case ◽

Tandem Repeats ◽

Direct Sequencing ◽

Single Cells ◽

Genetic Diagnosis ◽

Sex Selection ◽

Drop Out ◽

Gene Coding ◽

Custom Made

SummaryPreimplantation genetic diagnosis (PGD) of hemophilia A (HA) and other X-linked diseases through sex selection implies that male embryos will be systematically discarded, even though 50% are unaffected. The objective of the present work was to develop a PGD protocol for direct mutation identification that could be applied to first polar bodies (1PBs) in several HA clinical cases. Single buccal cells from controls and patients, and 1PBs were subjected to primer extension preamplification (PEP) PCR followed by amplification of F8 gene coding and intronic flanking regions, and direct sequencing. Moreover, multiplex fluorescent amplification of four short tandem repeats was adapted to a single cell preamplification in order to rule out contamination and allele drop-out, and for confirmatory indirect diagnosis. A couple at risk of HA transmission, with a familial mutation characterized as a 41-bp duplication in exon 14 of the F8 gene, was selected for the first clinical study. After optimizing the protocol, the complete F8 gene coding sequence was obtained from single cells to demonstrate the sensitivity of our methodology although in any clinical case only the relevant region, not the whole gene, must be amplified. The woman enrolled in the first clinical case has completed the first in-vitro fertilization cycle, and seven oocytes were analyzed with concordant results by both linkage analysis and direct sequencing method. Only one oocyte, among those diagnosed as mutation free, developed to embryo at day 3. It was transferred but pregnancy was not achieved. This PGD procedure enables non-affected and noncarrier embryo selection in families with any point or smallrange mutation in the F8 gene, without the need for further custom-made modifications.

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ASPEK ETIK PEMILIHAN JENIS KELAMIN DALAM PROSES PRE-IMPLANTATION GENETIC DIAGNOSIS PADA REKAYASA REPRODUKSI IN VITRO FERTILITATION

Aktualita (Jurnal Hukum) ◽

10.29313/aktualita.v2i2.4986 ◽

2019 ◽

Vol 2 (2) ◽

pp. 473-487

Author(s):

Budi Santoso

Keyword(s):

Preimplantation Genetic Diagnosis ◽

Genetic Disorder ◽

Genetic Diagnosis ◽

Sex Selection ◽

In Vitro Fertilisation

Perkembangan ilmu pengetahuan dan teknologi begitu pesatnya, denganilmu yang dimiliki oleh manusia, sudah banyak masalah yang berhasil dipecahkan. Perkembangan teknologi tersebut menjangkau segala hal, termasuk bidang kesehatan. Etika kedokteran juga sangat berhubungan dengan hukum. Hampir di semua negara ada hukum yang secara khusus mengatur bagaimana dokter harus bertindak dalam perawatan pasien dan penelitian. Kemajuan ilmu pengetahuan dan teknologi medis memunculkan masalah etis baru yang tidak dapat dijawab oleh etika kedokteran tradisional. Bagi pasangan atau pribadi yang tidak bisa menjadi hamil secara alami ada berbagai teknik reproduksi dengan bantuan seperti inseminasi buatan dengan fertilisasi in vitro dan transfer embrio, yang mudah didapat di pelayanan kesehatan reproduksi. Fertilisasi in vitro atau pembuahan in vitro (bahasa Inggris: in vitro fertilisation, IVF), atau sering disebut bayi tabung, adalah suatu proses pembuahan sel telur oleh sel sperma di luar tubuh sang wanita: in vitro ("di dalam gelas kaca"). Melalui teknologi preimplantation genetic diagnosis (PGD), tak hanya penyakit keturunan bisa dieliminasi, tapi jenis kelamin janin pun dapat dipilih. Pemilihan jenis kelamin (sex-selection) merupakan salah satu bentuk pengaplikasian dari teknologi rekayasa genetika yang berkembang cukup pesat saat ini. Di sini akan muncul pertanyaan apakah etis seseorang (orang-tua) menentukan jenis kelamin orang lain (anaknya) dengan sengaja? Di balik keberhasilan program bayi tabung, terdapat pula banyak masalah moral dan etika. Banyak pihak ynag beranggapan bahwa penelitian bayi tabung bermain main dengan kehidupan manusia karena telah mencampuri proses sacral dari penciptaan manusia yang merupakan hak prerogative Tuhan yang pencipta. Teknologi bayi tabung memberikan peluang kepada para pasangan untuk dapat mengetahui jenis kelamin dan kelainan genetik yang mungkin terjadi pada embrio, sehingga dapat menghindari kemungkinan implantasi embrio cacat. Seleksi kelamin atas indikasi medis dengan tujuan menghindari terjadinya sex linked genetic disorder , misalnya penyakit hemophilia dapat dibenarkan. Namun untuk indikasi nonmedik masih terdapat perbedaan pendapat. Seleksi kelamin ini tentunya menimbulkan perdebatan dari segi hukum, etika, dan social. Untuk indikasi nonmedik ini, ada yang setuju dan ada yang tidak setuju dengan seleksi kelamin. Jika sex-selection diperbolehkan secara bebas, sex-selection hanya akan menjadi industrialisasi di dunia kedokteran, karena akan menjadi semakin marak, dan chaos yang lebih banyak akan muncul.

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Ethical Issues and Practical Problems in Preimplantation Genetic Diagnosis

The Journal of Law Medicine & Ethics ◽

10.1111/j.1748-720x.1998.tb01902.x ◽

1998 ◽

Vol 26 (1) ◽

pp. 17-28 ◽

Cited By ~ 21

Author(s):

Jeffrey R. Botkin

Keyword(s):

Preimplantation Genetic Diagnosis ◽

Ethical Issues ◽

Chromosomal Abnormalities ◽

Genetic Diagnosis ◽

Embryonic Cells ◽

Cell Stage ◽

Vitro Fertilization ◽

Genetic Abnormalities ◽

Cellular Dna

Preimplantation genetic diagnosis (PGD) is a new method of prenatal diagnosis that is developing from a union of in vitro fertilization (IVF) technology and molecular biology. Briefly stated, PGD involves the creation of several embryos in vitro from the eggs and sperm of an interested couple. The embryos are permitted to develop to a 6-to-10-cell stage, at which point one of the embryonic cells is removed from each embryo and the cellular DNA is analyzed for chromosomal abnormalities or genetic mutations. An embryo or several embryos found to be free of genetic abnormalities are subsequently transferred to the woman's uterus for gestation. Embryos found to carry a genetic abnormality are discarded or frozen. Extra normal embryos may be frozen for future transfer or donation to another couple.

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Problems and prerequisites for determining the legal regime preimplantation genetic diagnosis in Russian legislation

10.21516/2413-1458-2019-18-2-175-181 ◽

2019 ◽

Author(s):

N.A. Altinnik , S.S. Zenin , V.V. Komarova et all ,

Keyword(s):

In Vitro Fertilization ◽

Preimplantation Genetic Diagnosis ◽

Human Embryo ◽

Legal Status ◽

Genetic Diagnosis ◽

Legal Regime ◽

Vitro Fertilization

Сurrent problems and prerequisites for the formation of the legal regime of pre-implantation genetic diagnosis (PGD) are considered in Russian legislation with account the existing approaches to determining the legal status of a “pre-implantation” embryo obtained in the framework of the in vitro fertilization procedure (IVF) are discussed. The authors substantiates the conclusion that it is necessary to legally determine PGD as one of the stages of using IVF, as well as establishing generally binding requirements for the procedure, conditions and features of this diagnosis, taking into account the need to minimize the damage caused to the human embryo.

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