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Quantitative evaluation by minisequencing and microarrays reveals accurate multiplexed SNP genotyping of whole genome amplified DNA
Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Sciences.
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2003 In: Nucleic Acids Research, Vol. 31, no 21, e129- p.Article in journal (Refereed) Published
Place, publisher, year, edition, pages
2003. Vol. 31, no 21, e129- p.
URN: urn:nbn:se:uu:diva-92069OAI: oai:DiVA.org:uu-92069DiVA: diva2:165021
Available from: 2004-09-15 Created: 2004-09-15Bibliographically approved
In thesis
1. Methods for Analysis of Disease Associated Genomic Sequence Variation
Open this publication in new window or tab >>Methods for Analysis of Disease Associated Genomic Sequence Variation
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

In Molecular Medicine a wide range of methods are applied to analyze the genome to find genetic predictors of human disease. Apart from predisposing disease, genetic variations may also serve as genetic markers in the search for factors underlying complex diseases. Additionally, they provide a means to distinguish between species, analyze evolutionary relationships and subdivide species into strains.

The development and improvement of laboratory techniques and computational methods was a spin-off effect of the Human Genome Project. The same techniques for analyzing genomic sequence variations may be used independent of organism or source of DNA or RNA. In this thesis, methods for high-throughput analysis of sequence variations were developed, evaluated and applied.

The performance of several genotyping assays were investigated prior to genotyping 4000 samples in a co-operative genetic epidemiological study. Sequence variations in the estrogen receptor alpha gene were found to be associated with an increased risk of breast and endometrial cancer in Swedish women.

Whole genome amplification (WGA) enables large scale genetic analysis of sparse amounts of biobanked DNA samples. The performance of two WGA methods was evaluated using four-color minisequencing on tag-arrays. Our in-house developed assay and “array of arrays” format allow up to 80 samples to be analyzed in parallel on a single microscope slide. Multiple displacement amplification by the Φ29 DNA polymerase gave essentially identical genotyping results as genomic DNA. To facilitate accurate method comparisons, a cluster quality assessment approach was established and applied to assess the performance of four commercially available DNA polymerases in the tag-array minisequencing assay.

A microarray method for genotyping human group A rotavirus (HRV) was developed and applied to an epidemiological survey of infectious HRV strains in Nicaragua. The method combines specific capture of amplified viral sequences on microarrays with genotype-specific DNA-polymerase mediated extension of capture oligonucleotides with fluorescent dNTPs.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2004. 89 p.
Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 0282-7476 ; 1371
Molecular medicine, microarray, molecular medicine, single nucleotide polymorphism, whole genome amplification, breast cancer, endometrial cancer, human rotavirus, Molekylärmedicin
National Category
Medical Genetics
urn:nbn:se:uu:diva-4525 (URN)91-554-6027-5 (ISBN)
Public defence
2004-10-08, Rudbecksalen, Rudbecklaboratoriet, Dag Hammarskjölds väg 20, Uppsala, 09:15
Available from: 2004-09-15 Created: 2004-09-15Bibliographically approved

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