Optimization of Single-Stranded Conformation Polymorphism (SSCP) Analysis for Screening for the Estrogen Receptor-α Gene Polymorphism P325P
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Darja Bitenc
Abstract
Since there are no theoretical models for single-stranded conformation polymorphism (SSCP) analysis, conditions for detecting specific mutation must be found experimentally. Previously, a time-consuming (22 hours) SSCP method was used for the detection of polymorphism in codon 325 (CCC to CCG; P325P) in exon 4 of estrogen receptor-α gene. The aim of our work was to study different gel loading buffers, additives to polyacrylamide gel, voltages, running times and temperatures of electrophoresis, in order to develop a better and faster SSCP analysis for screening of P325P polymorphism. Our results show that a low ionic strength gel loading buffer and 10% addition of glycerol to the 8% polyacrylamide gel (37:1) are essential for the good separation of mutated and wild-type single stranded conformers of exon 4. The most suitable conditions for electrophoresis were 300 V, 5 h and 22 °C. We concluded that a much faster SSCP analysis for sreening of P325P polymorphism of estrogen receptor-α gene was developed. Although our final result could be applied only to the detection of the described genetic polymorphism, we hope that the results of our study will be helpful to analysts using SSCP analysis in their mutation screening programs.
Copyright (c) 2001 by Walter de Gruyter GmbH & Co. KG
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- The Impact of Cardiac Natriuretic Peptide Determination on the Diagnosis and Management of Heart Failure
- Models for Combining Random and Systematic Errors. Assumptions and Consequences for different Models
- Nucleosomes in Serum as a Marker for Cell Death
- Comparative Nuclear Magnetic Resonance Studies of Water Permeability of Red Blood Cells from Maternal Venous Blood and Newborn Umbilical Cord Blood
- Optimization of Single-Stranded Conformation Polymorphism (SSCP) Analysis for Screening for the Estrogen Receptor-α Gene Polymorphism P325P
- Serum Total Renin after Tubal Sterilization
- Oxidative Stress and Male IGF-1, Gonadotropin and Related Hormones in Diabetic Patients
- New PCR-Based Method for the Sp1 Site Polymorphism in the COL1A1 Gene
- Comparability of Serum and Plasma Concentrations of Haemostasis Activation Markers
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- Arterial Tissue of Arsenic, Selenium and Iron in Blackfoot Disease Patients
- Evaluation of Aution Max AX-4280 Automated Urine Test-Strip Analyser
- Evaluation of Automated Enzyme Immunoassays for the Detection of Antibodies to Extractable Nuclear Antigens
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