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UBC Theses and Dissertations

Restriction endonuclease banding of human metaphase chromosomes Makihara, David Hajime


Restriction endonuclease banding is a recently introduced cytogenetic technique which reveals chromosomal polymorphisms at the centromeric region. These polymorphisms appear as variable sized structures and can be used in chromosome tracing analysis. In this project, metaphase chromosomes from normal subjects were banded using restriction endonuclease to establish the frequency of chromosomal polymorphism in the general population and also metaphase chromosomes were banded using restriction endonucleases in an attempt to trace the parental origin of embryonic chromosomes. The purpose of chromosome tracing analysis was to determine if uniparental disomy can be detected in early diploid spontaneous abortuses. The restriction endonuclease AluI was selected over RsaI, MboI, and DdeI, as it revealed reproducible polymorphisms at the centromeric regions of 20 out of 24 human chromosomes. The method of chromosome structure as standards was selected over the methods of linear measurements, surface area measurements, and polymorphism sizing, to quantify the size of the polymorphic region on each of the chromosomes. Using the chromosome structure as standards method, the frequencies of polymorphisms for each chromosome was calculated. This revealed that chromosomes 1,6,16, and Y were the most likely to be variable at the centromeric region. When parent to progeny chromosome tracing analysis was applied to metaphases treated with AluI restriction endonuclease, a visual method of tracing was superior to the method of chromosome structure as standards. Tracing analysis of Alul restriction endonuclease treated chromosomes revealed that 6% of chromosomes could be traced.

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