Simple and Easy to Perform Preimplantation Genetic Diagnosis for β-thalassemia Major Using Combination of Conventional and Fluorescent Polymerase Chain Reaction

Salehi, Rasoul; Khosravi, Sharifeh; Salehi, Mansour; Kheirollahi, Majid; Khanahmad, Hossein

Simple and Easy to Perform Preimplantation Genetic Diagnosis for β-thalassemia Major Using Combination of Conventional and Fluorescent Polymerase Chain Reaction

Document Type : Original Article

Authors

Department of Genetics and Molecular Biology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran

Abstract

Background: Thalassemias are the most common monogenic disorders in many countries throughout the world. The best practice to control the prevalence of the disease is prenatal diagnosis (PND) services. Extensive practicing of PND proved effective in reducing new cases but on the other side of this success high abortion rate is hided, which ethically unfair and for many couples, especially with a previous experience of a therapeutic abortion, or moral concerns, is not a suitable choice. Preimplantation genetic diagnosis (PGD) is a strong alternative to conventional PND. At present PGD is the only abortion free fetal diagnostic process. Considering the fact that there are more than 6000 single gene disorders affecting approximately 1 in 300 live-births, the medical need for PGD services is significant.
Materials and Methods: In the present study development of a PGD protocol for a thalassemia trait couple using nested multiplex fluorescent polymerase chain reaction (PCR) for the combination of polymorphic linked short tandem repeat (STR) markers and thalassemia mutations is described. Restriction fragment length polymorphism used to discriminate between wild and mutated alleles.
Results: In PGD clinical cycle, paternal and maternal alleles for D11S988 and D11S1338 STR markers were segregated as it was expected. PCR product for IVSII-1 mutation was subsequently digested with BtscI restriction enzyme to differentiate normal allele from the mutant allele. The mother's mutation, being a comparatively large deletion, was detectable through size differences on agarose gel.
Conclusion: The optimized single cell protocol developed and evaluated in this study is a feasible approach for preimplantation diagnosis of β-thalassemia in our patients.

Keywords

References

1.	Williams TN, Weatherall DJ. World distribution, population genetics, and health burden of the hemoglobinopathies. Cold Spring Harb Perspect Med 2012;2:a011692.
2.	Hardison RC, Chui DH, Riemer CR, Miller W, Carver MF, Molchanova TP, et al. Access to a syllabus of human hemoglobin variants (1996) via the World Wide Web. Hemoglobin 1998;22:113-27.
3.	Olivieri NF. The beta-thalassemias. N Engl J Med 1999 8;341:99-109.
4.	Abolghasemi H, Amid A, Zeinali S, Radfar MH, Eshghi P, Rahiminejad MS, et al. Thalassemia in Iran: Epidemiology, prevention, and management. J Pediatr Hematol Oncol 2007;29:233-8.
5.	Liao C, Mo QH, Li J, Li LY, Huang YN, Hua L, et al. Carrier screening for alpha-and beta-thalassemia in pregnancy: The results of an 11-year prospective program in Guangzhou Maternal and Neonatal hospital. Prenat Diagn 2005;25:163-71.
6.	Najmabadi H, Ghamari A, Sahebjam F, Kariminejad R, Hadavi V, Khatibi T, et al. Fourteen-year experience of prenatal diagnosis of thalassemia in Iran. Community Genet 2006;9:93-7.
7.	Järvholmab S, Brobergb M, Thurin-Kjellberga A. The choice of pre-implantation genetic diagnosis (PGD), a qualitative study among men and women. J Reprod Infant Psychol 2014;32:57-69.
8.	Kanavakis E, Traeger-Synodinos J. Preimplantation genetic diagnosis in clinical practice. J Med Genet 2002;39:6-11.
9.	Wells D, Sherlock JK. Strategies for preimplantation genetic diagnosis of single gene disorders by DNA amplification. Prenat Diagn 1998;18:1389-401.
10.	Sermon K. Current concepts in preimplantation genetic diagnosis (PGD): A molecular biologist's view. Hum Reprod Update 2002-Feb; 8:11-20.
11.	Ray PF, Handyside AH. Increasing the denaturation temperature during the first cycles of amplification reduces allele dropout from single cells for preimplantation genetic diagnosis. Mol Hum Reprod 1996;2:213-8.
12.	El-Hashemite N, Delhanty JD. A technique for eliminating allele specific amplification failure during DNA amplification of heterozygous cells for preimplantation diagnosis. Mol Hum Reprod 1997;3:975-8.
13.	Thornhill AR, McGrath JA, Eady RA, Braude PR, Handyside AH. A comparison of different lysis buffers to assess allele dropout from single cells for preimplantation genetic diagnosis. Prenat Diagn 2001;21:490-7.
14.	Vrettou C, Palmer G, Kanavakis E, Tzetis M, Antoniadi T, Mastrominas M, et al. A widely applicable strategy for single cell genotyping of beta-thalassaemia mutations using DGGE analysis: Application to preimplantation genetic diagnosis. Prenat Diagn 1999;19:1209-16.
15.	Findlay I, Ray P, Quirke P, Rutherford A, Lilford R. Allelic drop-out and preferential amplification in single cells and human blastomeres: Implications for preimplantation diagnosis of sex and cystic fibrosis. Hum Reprod 1995;10:1609-18.
16.	Findlay I, Quirke P, Hall J, Rutherford A. Fluorescent PCR: A new technique for PGD of sex and single-gene defects. J Assist Reprod Genet 1996;13:96-103.
17.	Van de Velde H, De Vos A, Joris H, Nagy ZP, Van Steirteghem AC. Effect of timing of oocyte denudation and micro-injection on survival, fertilization and embryo quality after intracytoplasmic sperm injection. Hum Reprod 1998;13:3160-4.
18.	Kolibianakis EM, Zikopoulos K, Verpoest W, Camus M, Joris H, Van Steirteghem AC, et al. Should we advise patients undergoing IVF to start a cycle leading to a day 3 or a day 5 transfer? Hum Reprod 2004;19:2550-4.
19.	Van Landuyt L, De Vos A, Joris H, Verheyen G, Devroey P, Van Steirteghem A. Blastocyst formation in in vitro fertilization versus intracytoplasmic sperm injection cycles: Influence of the fertilization procedure. Fertil Steril 2005;83:1397-403.
20.	Liebaers I, Sermon K, Staessen C, Joris H, Lissens W, Van Assche E, et al. Clinical experience with preimplantation genetic diagnosis and intracytoplasmic sperm injection. Hum Reprod 1998;13 Suppl 1:186-95.
21.	Kuliev A, Pomerantseva E, Polling D, Verlinsky O, Rechitsky S. PGD for inherited cardiac diseases. Reprod Biomed Online 2012;24:443-53.
22.	Quinn GP, Vadaparampil ST, Bower B, Friedman S, Keefe DL. Decisions and ethical issues among BRCA carriers and the use of preimplantation genetic diagnosis. Minerva Med 2009;100:371-83.
23.	SenGupta SB, Vadaparampil ST, Menon U. Preimplantation genetic diagnosis for hereditary cancers. Adv Exp Med Biol 2012;732:103-13.
24.	Noble R, Bahadur G, Iqbal M, Sanyal A. Pandora's box: Ethics of PGD for inherited risk of late-onset disorders. Reprod Biomed Online 2008;17 Suppl 3:55-60.
25.	Alberola TM, Vendrell X, Bautista-Llácer R, Vila M, Calatayud C, Pérez-Alonso M. Successful application of preimplantation genetic diagnosis for hypokalaemic periodic paralysis. Reprod Biomed Online 2010;21:206-11.
26.	Altarescu G, Brooks B, Eldar-Geva T, Margalioth EJ, Singer A, Levy-Lahad E, et al. Polar body-based preimplantation genetic diagnosis for N-acetylglutamate synthase deficiency. Fetal Diagn Ther 2008;24:170-6.
27.	Wilton L, Thornhill A, Traeger-Synodinos J, Sermon KD, Harper JC. The causes of misdiagnosis and adverse outcomes in PGD. Hum Reprod 2009;24:1221-8.
28.	Harper JC, Sengupta SB. Preimplantation genetic diagnosis: State of the art 2011. Hum Genet 2012;131:175-86.
29.	Hussey ND, Davis T, Hall JR, Barry MF, Draper R, Norman RJ, et al. Preimplantation genetic diagnosis for beta-thalassaemia using sequencing of single cell PCR products to detect mutations and polymorphic loci. Mol Hum Reprod 2002;8:1136-43.
30.	Shen X, Xu Y, Zhong Y, Zhou C, Zeng Y, Zhuang G, et al. Preimplantation genetic diagnosis for a-and ß-double thalassemia. J Assist Reprod Genet 2011;28:957-64.
31.	Langmore JP. Rubicon Genomics, Inc. Pharmacogenomics 2002;3:557-60.
32.	Fiorentino F, Magli MC, Podini D, Ferraretti AP, Nuccitelli A, Vitale N, et al. The minisequencing method: An alternative strategy for preimplantation genetic diagnosis of single gene disorders. Mol Hum Reprod 2003;9:399-410.
33.	Vanneste E, Melotte C, Voet T, Robberecht C, Debrock S, Pexsters A, et al. PGD for a complex chromosomal rearrangement by array comparative genomic hybridization. Hum Reprod 2011;26:941-9.
34.	Handyside AH, Harton GL, Mariani B, Thornhill AR, Affara N, Shaw MA, et al. Karyomapping: A universal method for genome wide analysis of genetic disease based on mapping crossovers between parental haplotypes. J Med Genet 2010;47:651-8.
35.	De Rycke M, Van de Velde H, Sermon K, Lissens W, De Vos A, Vandervorst M, et al. Preimplantation genetic diagnosis for sickle-cell anemia and for beta-thalassemia. Prenat Diagn 2001;21:214-22.
36.	Figueira RC, Setti AS, Cortezzi SS, Martinhago CD, Braga DP, Iaconelli A, Jr. et al. Preimplantation diagnosis for beta-thalassemia combined with HLA matching:First “savior sibling” is born after embryo selection in Brazil. J Assist Reprod Genet 2012;29:1305-9.
37.	Van de Velde H, De Rycke M, De Man C, De Hauwere K, Fiorentino F, Kahraman S, et al. The experience of two European preimplantation genetic diagnosis centres on human leukocyte antigen typing. Hum Reprod 2009;24:732-40.

Advanced Biomedical Research