[Frontiers in Bioscience E4, 1049-1057, January 1, 2012]

LOH of PTCH1 region in BCC and ovarian carcinoma: microsatellite vs. HRM analysis

Vesna Musani1, Maja Sabol1, Diana Car1, Petar Ozretic1, Slavko Oreskovic2, Dinko Leovic3, Sonja Levanat1

1Division of Molecular Medicine, Rudjer Boskovic Institute, Zagreb, Croatia, 2Department of Obstetrics and Gynecology, University of Zagreb School of Medicine, Zagreb, Croatia, 3Department of Maxillofacial Surgery, University Hospital Osijek, Osijek, Croatia

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Materials and methods
3.1. Patient samples and DNA extraction
3.2. Microsatellite analysis of LOH
3.3. High resolution melting analysis of SNPs
3.4. DNA sequencing
3.5. Statistical analysis
4. Results
4.1. Microsatellite analysis
4.2. HRM analysis
4.3. Comparison of two methods
5. Discussion
6. Acknowledgements
7. References

1. ABSTRACT

Loss of heterozygosity (LOH) of tumor suppressor genes is a frequent event in tumorigenesis. LOH is most often analyzed by microsatellite typing, but here we offer a fast and efficient method for simultaneous SNP genotyping and mutation scanning, which can also be used for LOH detection. High resolution melting (HRM) provides simple variant detection, and can be adopted for a wide range of applications. When a melting profile for a specific SNP is determined, the screening can be done without the need for sequencing, and only the melting profiles differing from the established melting profiles should be sequenced. LOH of PTCH1 gene is often found in a series of different tumors, for example basal cell carcinoma (BCC) and ovarian carcinoma (OC). In this study LOH was detected in 50% of BCC and 27.27% of OC, and the detection rates of microsatellite typing and HRM were comparable. Both methods depended only on the heterozygosity of the loci analyzed, but HRM offers an additional advantage of detection of all sequence variants in the gene of interest.