Quoc Ty Tran1,2,3, Tatjana Jatsenko1, Olev Poolamets4, Olga Tšuiko1, Dmitri Lubenets5, Tiia Reimand2,6, Margus Punab4, Maire Peters7,8, Andres Salumets1,2,9,10. 1. Competence Centre on Health Technologies, Tiigi 61B, 50410, Tartu, Estonia. 2. Institute of Biomedicine and Translational Medicine, University of Tartu, Tartu, Estonia. 3. Danang University of Medical Technology and Pharmacy, Da Nang, Vietnam. 4. Andrology Centre, Tartu University Hospital, Tartu, Estonia. 5. Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia. 6. Department of Clinical Genetics, Tartu University Hospital, Tartu, Estonia. 7. Competence Centre on Health Technologies, Tiigi 61B, 50410, Tartu, Estonia. maire.peters@ut.ee. 8. Department of Obstetrics and Gynaecology, Institute of Clinical Medicine, University of Tartu, Tartu, Estonia. maire.peters@ut.ee. 9. Department of Obstetrics and Gynaecology, Institute of Clinical Medicine, University of Tartu, Tartu, Estonia. 10. Department of Obstetrics and Gynecology, University of Helsinki and Helsinki University Hospital, Helsinki, Finland.
Abstract
PURPOSE: The purpose of this study was to develop a feasible approach for single sperm isolation and chromosome analysis by next-generation sequencing (NGS). METHODS: Single sperm cells were isolated from semen samples of normozoospermic male and an infertile reciprocal translocation (RcT) carrier with the 46,XY,t(7;13)(p12;q12.1) karyotype using the optimized fluorescence-activated cell sorting (FACS) technique. Genome profiling was performed using NGS. RESULTS: Following whole-genome amplification, NGS, and quality control, the final chromosome analysis was performed on 31 and 6 single cell samples derived from the RcT carrier and normozoospermic male, respectively. All sperm cells from normozoospermic male showed a normal haploid 23-chromosome profile. For the RcT carrier, the sequencing data revealed that 64.5% of sperm cells harbored different variants of chromosome aberrations, involving deletion of 7p or 7q, duplication of 7p, and duplication of 13q, which is concordant with the expected chromosome segregation patterns observed in balanced translocation carriers. In one sample, a duplication of 9q was also detected. CONCLUSIONS: We optimized FACS protocol for simple and efficient isolation of single human sperm cells that subsequently enabled a successful genome-wide chromosome profiling and identification of segmental aneuploidies from these individual cells, following NGS analysis. This approach may be useful for analyzing semen samples of infertile men or chromosomal aberration carriers to facilitate the reproductive risk assessment.
PURPOSE: The purpose of this study was to develop a feasible approach for single sperm isolation and chromosome analysis by next-generation sequencing (NGS). METHODS: Single sperm cells were isolated from semen samples of normozoospermic male and an infertile reciprocal translocation (RcT) carrier with the 46,XY,t(7;13)(p12;q12.1) karyotype using the optimized fluorescence-activated cell sorting (FACS) technique. Genome profiling was performed using NGS. RESULTS: Following whole-genome amplification, NGS, and quality control, the final chromosome analysis was performed on 31 and 6 single cell samples derived from the RcT carrier and normozoospermic male, respectively. All sperm cells from normozoospermic male showed a normal haploid 23-chromosome profile. For the RcT carrier, the sequencing data revealed that 64.5% of sperm cells harbored different variants of chromosome aberrations, involving deletion of 7p or 7q, duplication of 7p, and duplication of 13q, which is concordant with the expected chromosome segregation patterns observed in balanced translocation carriers. In one sample, a duplication of 9q was also detected. CONCLUSIONS: We optimized FACS protocol for simple and efficient isolation of single human sperm cells that subsequently enabled a successful genome-wide chromosome profiling and identification of segmental aneuploidies from these individual cells, following NGS analysis. This approach may be useful for analyzing semen samples of infertilemen or chromosomal aberration carriers to facilitate the reproductive risk assessment.
Authors: Olga Tšuiko; Tuuli Dmitrijeva; Katrin Kask; Pille Tammur; Neeme Tõnisson; Andres Salumets; Tatjana Jatsenko Journal: Mol Cytogenet Date: 2019-06-18 Impact factor: 2.009
Authors: Roger W L Godschalk; Carole L Yauk; Jan van Benthem; George R Douglas; Francesco Marchetti Journal: Environ Mol Mutagen Date: 2019-12-05 Impact factor: 3.216