Elizabeth Normand1, Sadeem Qdaisat1, Weimin Bi1, Chad Shaw1, Ignatia Van den Veyver1,2, Arthur Beaudet1, Amy Breman3. 1. Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, USA. 2. Department of Obstetrics and Gynecology, Baylor College of Medicine, Houston, TX, USA. 3. Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, USA. breman@bcm.edu.
Abstract
OBJECTIVE: Detection of genomic copy number abnormalities in a single cell using array comparative genomic hybridization (CGH) offers a promising non-invasive alternative for prenatal diagnosis. Our objective was to compare three commercially available whole-genome amplification (WGA) kits for their capacity to produce high quality DNA from single cells that is suitable for both molecular genotyping and array CGH. METHODS: We examined kit performance on unfixed, fixed and fixed/permeabilized lymphoblastoid cells. Molecular genotyping methods were used to evaluate the fidelity of amplified DNA for genomic profiling, while array CGH was used to assess copy number from single cells harboring trisomy 21, a DiGeorge syndrome deletion, a CMT1A duplication or a MECP2 duplication. RESULTS: Molecular genotyping was achieved from single cells but performance varied between WGA kits. Furthermore, we consistently detected a dosage difference in sex chromosomes for gender mismatched hybridizations and for chromosome 21 in trisomy 21 cells. The 2.5 Mb DiGeorge syndrome deletion was also detected using all three WGA platforms, whereas the 1.3 Mb CMT1A and the 0.6 Mb MECP2 duplications were not consistently detected. CONCLUSION: These data suggest that single cell molecular genotyping and copy number analysis can be accomplished when WGA conditions are optimized.
OBJECTIVE: Detection of genomic copy number abnormalities in a single cell using array comparative genomic hybridization (CGH) offers a promising non-invasive alternative for prenatal diagnosis. Our objective was to compare three commercially available whole-genome amplification (WGA) kits for their capacity to produce high quality DNA from single cells that is suitable for both molecular genotyping and array CGH. METHODS: We examined kit performance on unfixed, fixed and fixed/permeabilized lymphoblastoid cells. Molecular genotyping methods were used to evaluate the fidelity of amplified DNA for genomic profiling, while array CGH was used to assess copy number from single cells harboring trisomy 21, a DiGeorge syndrome deletion, a CMT1A duplication or a MECP2 duplication. RESULTS: Molecular genotyping was achieved from single cells but performance varied between WGA kits. Furthermore, we consistently detected a dosage difference in sex chromosomes for gender mismatched hybridizations and for chromosome 21 in trisomy 21 cells. The 2.5 Mb DiGeorge syndrome deletion was also detected using all three WGA platforms, whereas the 1.3 Mb CMT1A and the 0.6 Mb MECP2 duplications were not consistently detected. CONCLUSION: These data suggest that single cell molecular genotyping and copy number analysis can be accomplished when WGA conditions are optimized.
Authors: Andrea D Tyler; Laura Mataseje; Chantel J Urfano; Lisa Schmidt; Kym S Antonation; Michael R Mulvey; Cindi R Corbett Journal: Sci Rep Date: 2018-07-19 Impact factor: 4.379