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The PCR method optimization for analysis of the gene PAI-1 polymorphism
SPĚVÁKOVÁ, Daniela
The plasminogen activator inhibitor (PAI-1) is an important biomarker of fibrinolysis. This glycoprotein acts as a principal inhibitor of plasminogen activators and its high plasma level is associated with various types of diseases, such as cardiovascular diseases or malignant tumors. The PAI-1 gene is located on chromosome 7 in region 7q22.1. In 1993 4G/5G polymorphism was discovered in the gene promoter. The 4G allele increases PAI-1 gene expression which results in a higher plasma level of this inhibitor. That's why this polymorphism has become the subject of many studies. The main aim of my bachelor thesis was to optimize the ARMS-PCR method for analysis of the gene PAI-1 4G/5G polymorphism. Other goals were to write a specialized research on this subject and to learn the techniques used in the genetic laboratory (DNA isolation, DNA concentration measurement, Real-time PCR). In the theoretical part I describe the PAI-1 gene and its product. I deal mainly with the role of this inhibitor in the mechanism of fibrinolysis and its influence on various diseases. Furthermore, I showed an overview of methodological approaches used for genetic analysis of 4G/5G polymorphism in PAI-1 gene. In the practical part there are presented procedures of optimization of the ARMS PCR method and the results of the genotypes of the examined group obtained by the Real-time PCR method, when a commercial kit was used. The optimization of the ARMS-PCR method was not successful.
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