Successful Strategy of Pre-implantation Genetic Testing for Beta-Thalassemia (c.17A>T Mutation)-Hb E Disease Using Multiplex Fluorescent PCR and Mini-Sequencing
Worashorn Lattiwongsakorn, Natpat Jansaka, S. Piyamongkol, T. Pantasri, T. Tongsong, Wanwisa Suriya, W. Piyamongkol
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引用次数: 0
Abstract
Objectives: Hemoglobin E disease, c.26G>A variant of beta-globin gene, is the most common hemoglobinopathy in Asia. Compound heterozygotes inheriting Hb E disease and beta-thalassemia generate beta-thalassemia-Hb E disease with severe anemia. This study aimed to develop a pre-implantation genetic testing for monogenic disorders (PGT-M) protocol for beta–thalassemia (c.17A>T mutation)-Hb E disease (c.26G>A mutation) using multiplex fluorescent polymerase chain reaction (PCR) and mini-sequencing. Materials and Methods: bthalw1 primers were used to amplify a beta-globin gene fragment covering both mutations, i.e. beta– thalassemia (c.17A>T) and Hb E disease. D21S11 microsatellite marker was included for contamination detection. Novel mini-sequencing primers were designed and tested for detection of both mutations. Results: Pre-clinical work up of the optimized PGT-M protocol using 20 single buccal cells of a heterozygous subject showed 100% amplification efficiency and 0% allele drop out (ADO) rate for both primers. In clinical PGT-M cycle, 15 embryos were subjected to biopsy. The results showed two normal, one heterozygous for beta-thalassemia, six heterozygous for Hb E disease, one affected for beta-thalassemia-Hb E disease and five with ambiguous results. Two normally diagnosed embryos were chosen for transfer, one singleton pregnancy was obtained. A healthy baby boy was resulted. Postnatal testing confirmed PGT results. Conclusions: Novel PGT-M protocols for beta-thalassemia-Hb E disease using multiplex fluorescent PCR and mini-sequencing were developed and described here. The protocol was applied in a clinical PGT-M cycle and gave rise to one successful pregnancy and consequently a healthy baby boy. Mini-sequencing was proved to be rapid, accurate and cost-effective protocol for PGT-M.
期刊介绍:
All kind of knowledge contributing to the development of science by its content, value, level and originality will be covered by IJWHR. Problems of public health and their solutions are at the head of the windows opening us to the world. The "International Journal of Women''s Health and Reproduction Sciences” is a modern forum for scientific communication, covering all aspects women health and reproduction sciences, in basic and clinical sciences, mainly including: -Medical Education in Women Health and Reproduction Sciences -Cardiology in Women Health-Related Reproductive Problems -Sports Medicine in Women Health and Reproduction Sciences -Psychiatry in Women Health-Related Reproductive Problems -Antioxidant Therapy in Reproduction Medicine Sciences -Nutrition in Women Health and Reproduction Sciences -Defense Androgen and Estrogen -Fertility and Infertility -Urogynecology -Endometriosis -Endocrinology -Breast Cancer -Menopause -Puberty -Eroticism -Pregnancy -Preterm Birth -Vaginal Diseases -Sex-Based Biology -Surgical Procedures -Nursing in Pregnancy -Obstetrics/Gynecology -Polycystic Ovary Syndrome -Hyperandrogenism in Females -Menstrual Syndrome and Complications -Oncology of Female Reproductive Organs -Traditional Medicine in Women Reproductive Health -Ultrasound in Women Health Reproduction sciences -Stem Cell Research In Women Reproduction Sciences -Complementary Medicine in Women Reproductive Health -Female Sexual Dysfunction: Pathophysiology & Treatment