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dc.description.abstract본 실험실에서는 위암과 간암, 애기장대 조직의 유전자 발현을 조사하기 위해 SAGE, SSH, GLGI, cDNA microarray 실험을 수행하였다. 본 논문에서는 이들 실험으로부터 발생된, 서로 연관성을 지니고 있는 대량의 실험 결과들을 효율적으로 활용하기 위한 데이터베이스 구축과 이를 활용한 SSH technique의 효율성 분석에 대한 내용을 다루고 있다. 각 조직에 대한 실험결과를 통합하고 있는 데이터베이스 시스템은 웹 상의 사용자 페이지를 통해 접근이 가능하다. ( 각 실험들은 하나의 조직에 대해 여러 실험이 수행되고 한 실험이 여러 조직에 대해 수행된 다(多) 대 다(多) 관계를 가지고 있기 때문에, 이들 실험 결과의 비교 분석을 통해 실험의 효율성이나 더욱 정확한 세포내의 유전자 발현양상을 알 수 있다. 본 논문에서 실험적 효율성을 확인해보고자 하는 SSH technique은 일반적으로 suppression PCR 효과에 의해 세포 내의 mRNA 발현 빈도 차이 문제를 극복한다. 따라서 차등적으로 발현되는 유전자 중에서 특히 낮은 발현양상을 나타내는 유전자를 연구하고자 할 때 많이 사용되는 기법이다. SSH 실험이 수행된 동일 조직에 대해 유전자 발현양상을 tag count 나 signal intensity 같은 구체적인 값으로 나타내주는 SAGE나 microarray 실험이 수행되었기 때문에 SSH 실험에서 얻은 구체적인 유전자의 발현수준을 확인 해 볼 수 있었다. SAGE나 microarray 유전자 발현 profiles에서 발견되는 SSH 유전자의 대부분은 낮은 발현 수준을 보이는 그룹에 속했다. 즉 SSH 실험을 통해 얻은 유전자에는 낮은 발현수준을 보이는 유전자가 많다는 의미이며 이것은 SSH 가 낮은 발현양상을 보이는 유전자를 연구하는데 적절한 실험 방법임을 증명한다. ;To identify and isolate differentially expressed transcripts, there are number of techniques available, such as differential display (DD), representational difference analysis (RDA), serial analysis of gene expression (SAGE), suppression subtractive hybridization (SSH) and cDNA microarray. One advantage of the SSH technique is the equalization of high and low abundance messages, which allows for the identification of differentially expressed low abundance mRNA. Despite its advantages to enrich and isolate differentially expressed genes, practical efficacy of the method has not been thoroughly studied by using quantitative method such as SAGE and microarray. In this study, SSH technique efficiency was verified by comparing expression level of the genes derived from SSH in expression profiles by SAGE and cDNA microarray experiments carried out on identical RNA specimens for three different sets of tissues, i.e. two sets of human gastric normal/cancer tissues and a set of Arabidopsis normal/cold-treated leaf. Result of the investigation into the tag count for the genes from SSH overlapped in SAGE profiles of the human gastric cancer reveals that over 90% of the genes had below 10 tag count, while, proportion of the genes to have below 10 tag count in Arabidopsis cold treated leaf was 83.2%. This lower transcription level of the genes from SSH was appeared also in the analysis of hybridization intensities in microarray. Respectively, 87% of the genes from human gastric cancer and Arabidopsis cold treated leaf belong to the group with lower intensity than average of intensity measurements. In both SAGE and microaray experiments, many genes observed in the repressed genes from SSH had low fold differences. Especially, proportion of the genes representing low fold differences in microarray was much lower than in SAGE. The feature of the genes derived from SSH suggests that many genes to have low transcription level were included in the genes obtained in the SSH experiment and demonstrate that SSH technique is reasonably reliable for obtaining low abundance transcripts.-
dc.description.tableofcontentsLIST OF FIGURES = iii LIST OF TABLES = v CHAPTER I. Construction of in-house databases for cancer and cold stress research = 1 INTRODUCTION = 2 IMPLEMENTATION = 9 DATABASE LAYOUT = 18 Human gastric cancer-related database (GC_DB) = 20 Arabidopsis cold stress-related database (CS_DB) = 22 Human liver cancer-related database(LC_DB) and Primer-related database(Primer_DB) = 24 WEB interface(Using the database system on web) = 27 Search page for gastric cancer-related information = 27 Search page for Arabidopsis cold stress-related information = 36 Search page for liver cancer-related information = 37 REFERENCES = 38 CHAPTER II. Verification of efficiency of SSH approach for identifying low abundance transcripts of differentially expressed genes = 40 ABSTRACT = 41 INTRODUCTION = 43 MATERIALS and METHODS = 48 RESULTS = 49 1. Sequence analysis of the clones generated by SSH = 50 2. Express pattern of the genes from SSH in the gene expression profile by SAGE = 54 3. Express pattern of the genes from SSH in the gene expression profile by cDNA microaray = 60 DISCUSSION = 63 REFERENCES = 68 국문초록 = 70-
dc.format.extent2952772 bytes-
dc.publisher이화여자대학교 대학원-
dc.titleConstruction of in-house databases and comparative verification of SSH approach-
dc.typeMaster's Thesis-
dc.format.pagev, 70 p.-
dc.identifier.major대학원 생명과학과- 2-
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