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KMID : 0384119970170020351
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Korean Journal of Clinical Pathology
1997 Volume.17 No. 2 p.351 ~ p.359
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Detection of N-myc Gene Amplification in Neuroblastoma Using the Semiquantitative Polymerase Chain Reaction
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박덕우
이창훈/윤갑준/김황민/양우익
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Abstract
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Background :
@EN The abnormality of N myc gene was found in human neuroblastomas, retinoblastomas and small-cell lung carcinomas, where it was often amplified or overexpressed. It was therefore suggested that the abnormal expression of N-myc gene might play a
role
in the development or progression of these tumors and poor prognosis. Conventional techniques for N-myc gene quantification by Southern hybridization, using radiolabeled probes, time consuming and labor intensive. We have used the polymerase
chain
reaction (PCR) to detect amplification of the N-myc oncogene in extracted DNA from cell-lines and paraffin-embedded sections.
@ES Methods :
@EN Twenty-four paraffin-embedded neuroblastoma tissues were evaluated for N-myc gene amplification. We used two primer pairs for co-amplifying the β-globin (internal control) and N- myc gene in the same PCR reaction tube. At this time, the
r12-P
dATP
was 5' end-labeled at each sense primers by T4 polynucleotide kinase method. Three neuroblastoma cell lines were used as external controls : LAN-6, IMR32, and KCNR cell line, After 30 cycles of -PCR, the products were resolved by 1.5% agarose gel
electrophoresis. N-myc gene amplification was identified by visual comparison of the relative intensities of β-globin product band on the ethidium-bromide-stained-gel. Amplified PCR prduct bands were quantitated by densitometry on polyacylamide
gel
autoradiographs.
@ES Results :
@EN By visual inspection, three of 24 cases were positive and four were indeterminant for N-myc gene amplification, respectively. By densitometry on autoradiographs, seven of 24 (29%) cases were positive including four indeterminant cases by
visual
inspections. And also. N-myc gene amplifications were detected in 2 of 5 (40%) cases in clinical stage II and III. respectively, and 3 of 9 (33%) cases in stage IV.
@ES Conclusions :
@EN We conclude that semiquantitative PCR is rapid and reliable method for quantitative alterating detection of N-myc gene amplification in neuroblastomas, and that can be performed rapidly on DNA extracted from paraffin-embedded tissue or frozen
section. (Korean J Clin Pathol 1997 : 17(2) : 351~9)
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