Solid-phase minisequencing confirmed by FISH analysis in determination of gene copy number |
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Authors: | Maris Laan Kristiina Grön-Virta Armi Salo Pertti Aula Leena Peltonen Aarno Palotie Ann-Christine Syvänen |
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Affiliation: | (1) Department of Clinical Chemistry, University of Helsinki and the Laboratory Department of Helsinki University Hospital, FIN-00290 Helsinki, Finland;(2) Department of Human Molecular Genetics, National Public Health Institute, FIN-00300 Helsinki, Finland;(3) Department of Medical Genetics, University of Turku, FIN-20520 Turku, Finland;(4) United Laboratories Ltd., P.O.Box 222, FIN-00381 Helsinki, Finland |
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Abstract: | The solid-phase minisequencing method (Syvänen et al. 1990) allows accurate quantative determination of the ratio between two DNA or RNA sequences that are present as a mixture in a sample and differ from each other only by a single nucleotide. Here, we present another application of the minisequening method, the determination of the gene copy number in a genome. The copy number of a marker gene aspartyl glucosaminidase (AGA) located at 4qter, was determined in three patients with a chromosomal alteration involving the distal region of 4q. For the minisequencing assay an equal amount of DNA from a patient homozygous for a mutation in the AGA gene was added to the DNA samples concerned. The relative amount of the normal sequence determined in each combined sample gives the copy number of the AGA gene. Fluorescence in situ hybridization (FISH), applied in parallel as a control, produced concordant results with solid-phase minisequencing in each case. As the potential of the minisequencing lies in automation, it could be a useful tool in the screening of monosomies, trisomies or loss of heterozygosity in diagnostics. |
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