Differential PCR 과 Magnetic Bead/단 클론 항체법을 이용한 암유전자 N-myc 의 증폭분석법 개발 및 N-myc 의 증폭과 발현에 관한 연구

다국어 초록 (Multilingual Abstract)

Analysis of risk factors at the diagnosis of cancer is essential for selecting appropriate therapy. Since genomic amplification of N-myc is an powerful prognostic indicator of neuroblastoma and its marrow metastases provide a ready source of tumor cells for fundamental and clinical studies, we have adapted the fluorescence-based detection system of the automated DNA sequencer to quantitate N-myc amplification in neuroblastoma cells that have metastasized to bone marrow. Fluorescence-labelled primer pairs were used to co-amplify a 428-bp fragment of N-myc along with a 268-bp fragment of β-globin gene(a single copy reference standard). After 30 cycles of PCR, the products were denatured by heating in formamide and the single-stranded products were resolved on polyacrylamide/urea gels. Signal quantitation and molecular sizing were performed to analyse N-mycl β-globin signal ratio. This assay was sufficiently senstitive to detect N-myc amplification from marrow specimen containing 2％ tumor cells without enrichment and showed a relatively high linear regression(R=0.88) indicating relationship between N-mycl β-globin ratio and percentage of tumor cells in bone marrow. Ficoll/Hypaque sedimentation, which enable 3.7-fold enrichment, followed by positive magnetic enrichment can concentrate tumor cells one and half logs. The differntial PCR combined with immunomagnetic enrichment was sensitive enough to detect N-myc amplification in the bone marrow containing around 0.1％ neuroblastoma cells.
These results suggest that this quantitative PCR, which has provided a simple, rapid, nonisotopic method using only small amounts of tissue, can be applied to prognosticate neuroblastoma patients by using clinical marrows when primary tumors are not available.

목차 (Table of Contents)

• 서론
• 실험재료 및 방법
• 결과
• 고찰
• 결론
• 참고문헌
• Figure Legends
• 발표초록
• 연구비 사용 내역
• 연구결과 활용방안
• 참고 및 건의사항
• 배출인력양성
• 요약문