A RAPID METHOD FOR THE DETECTION OF ALLELIC IMBALANCE OF THE LONG ARM OF CHROMOSOME 18 IN COLORECTAL CANCER
Stefanovska A-M1*, Jasar D2*, Zografski G2, Josifovski T3, Panovski M3, Efremov GD1, Dimovski AJ1
*Corresponding Author: Dr. Aleksandar J. Dimovski, Macedonian Academy of Sciences and Arts, Research Center for Genetic Engineering and Biotechnology, Aven. Krste Misirkov 2, POB 428,1000 Skopje, Republic of Macedonia; Tel: +3892-120253; Fax: +3892-115434; E-mail: aleks@amanu.edu.mk
page: 21

Abstract

Allelic loss or loss of heterozygosity (LOH) at 18q is a common event in colorectal cancerogenesis that indicates the loss of several tumor suppressor genes. It has been suggested that 18q allelic loss is an independent factor for relapse in colorectal cancer patients with Dukes' stages B and/or C. The conventional method for assessment of 18q allelic loss is based on PCR of several microsatellite markers, electrophoresis on denaturing long polyacrylamide gels, visualization of fragments by autoradiography and quantification of allelic content by densitometry or scintillation counting. We have developed a fast, easy and reproducible assay for 18q allelic loss assessment, which is based on fluorescent multiplex PCR of five microsatellite markers [D18S58, D18S61, DCC, D18S46 (DPC4) and D18S535] and fragment analysis by automated ABIPrism 310 Genetic analyzer. The loss of heterozygosity at a particular locus was scored when the allele ratio in the cancer differed by 35% from the ratio in the normal tissue. Loss of heterozygosity at 18q was present in 31 of 92 (34%) analysed cancers. The results obtained by the multiplex fluorescent PCR were in a complete agreement with the data obtained by conventional method for 18q allelic loss detection. Due to its simplicity, speed, reproducibility and lesser manipulation requirements, the automated fluorescent multiplex PCR analysis is a method of choice for a routine diagnosis of allelic imbalances of clinical relevance.




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