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研究生:許維中
研究生(外文):HSU WEI JONG
論文名稱:cDNA微陣列晶片上探針交互雜交問題之探討與分析cDNA微陣列晶片上探針交互雜交問題之探討與分析cDNA微陣列晶片上探針交互雜交問題之探討與分析
論文名稱(外文):Evaluation and Analysis of Cross Hybridization Effects of Probes on cDNA Microarrays
指導教授:江彥逸江彥逸引用關係
學位類別:碩士
校院名稱:長庚大學
系所名稱:資訊管理研究所
學門:電算機學門
學類:電算機一般學類
論文種類:學術論文
論文出版年:2005
畢業學年度:93
語文別:中文
中文關鍵詞:DNA微陣列技術cDNA微陣列交互雜交相似性序列BLAST
外文關鍵詞:DNA microarray technologycDNA microarrayCross hybridizationsimilar sequenceBLAST
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  • 被引用被引用:1
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DNA微陣列高產量的基因表現分析,近年來廣泛運用於觀察基因的表現。在DNA微陣列實驗中,原則上一個探針應該只會與一個相對應的基因雜交,但是基因序列之間的高度相似性、同源性,可能造成DNA微陣列實驗過程中探針交互雜交的現象,而交互雜交的現象會造成實驗結果判讀的誤差。
本研究即是針對cDNA微陣列上探針可能發生交互雜交的情形做探討,並以現行臨床使用的微陣列晶片作為觀察的目標。透過序列排比工具(BLAST)找尋與探針序列相似的基因序列,並且將這些相似性序列與探針的序列資訊整合,建立探針可能發生交互雜交的探針基因對應資料庫。
我們從資料庫中找尋可能發生交互雜交的探針,並由微陣列實驗結果所得到的數據資料,觀察相似基因探針組表現量的相關程度。從可能發生交互雜交成對的探針組中,其非目標基因的相似長度與表現量具有高度相關性。相似長度越長的探針組,其表現量也具有較高的關係。本研究建議當非目標基因相似長度大於300 bp且相似度超過80%以上時,在分析這些實驗數據應考慮交互雜交所帶來的影響。
Microarray have been extensively used to studying genome-wide expression analysis. A potential problem with microarray research is that probes of an array might have segments of similar sequence that resulted form gene families, homologous genes and highly similar sequences may cause cross hybridization. Cross hybridization effects can be a source of error and reducing the reliability of DNA microarray results.
The objective of this research is to discuss observations of cross hybridiztaion on an existing set of cDNA microarrays that are currently used by Chang Gung genomic center (CGG). We first submit the probes sequences to the NCBI database and use the BLAST algorithm to find the possible probe-to-gene mappings. We use this information to identify the possible cross-hybridizing paired probes on the array. We attempt to establish criteria that can influence or resulted in cross hybridizations on cDNA microarrays.We use statistical methods to evaluate the impact of cross hybridization on the selected sequence-matched probes. Our results have shown that there is a strong relationship between the intensity values and the match length. If the match length is sufficiently long , then we can observe a high correlation between the expression of the sequence -matched probes.
Results obtained in this exploratory study have suggested that if the non-target sequence have at least 300bp of contiguous matching length with similarity over 80% with respect to the cDNA probe, then researchers should consider the phenomenon of cross hybridization when interpreting expression profile. Based on this observation, we recommend some rules that can be followed when analysising CGG’s cDNA microarrays.
目 錄
指導教授推薦書……………………………………………………….
口試委員會審定書……………………………………………………
致謝……………………………………………………………………
中文摘要………………………………………………………………
英文摘要………………………………………………………………
第一章 緒論……………………………………………………………1
1-1 研究動機…………………………………………………………2
1-2 研究目的…………………………………………………………3
1-3 論文架構及研究流程……………………………………………5
第二章 文獻探討 …………………………………………………8
2-1 DNA 微陣列……………………………………………………8
2-1-1 DNA 微陣列的簡介………………………………………8
2-1-2 cDNA實驗過程……………………………………………10
2-1-3 探針的製作………………………………………………12
2-2 交互雜交的探討………………………………………………13
2-2-1 核酸的雜交………………………………………………13
2-2-2 過去交互雜交的研究……………………………………14
2-2-3 DNA微陣列上交互雜交的型態……………………………15
第三章 操作原理……………………………………………………17
3-1第一型交互雜交探針的尋找…………………………………17
3-1-1自動化BLAST比對相似性序列……………………………19
3-1-2 BLAST 比對結果…………………………………………20
3-1-3 連結Unigene擷取相似基因的資訊……………………23
3-2 第二型交互雜交探針的尋找…………………………………24
3-3 探針的資訊……………………………………………………24
3-4 程式開發與實作………………………………………………25
第四章 探針基因對應資料庫分析…………………………………26 4-1 cDNA微陣列晶片探針的資訊…………………………………26
4-2 探針基因對應資料庫的比對結果分析………………………28
4-3 決定每點探針應雜交的目標基因……………………………29
4-4 一個探針對到兩個基因的分析結果…………………………33
4-5 一個探針對到三個基因的分析結果…………………………34
4-6 找出可能的交互雜交的探針…………………………………36
4-6-1第一型交互雜交探針的尋找……………………………34
4-6-2第二型交互雜交探針的尋找……………………………37
第五章 結果…………………………………………………………38
5-1 微陣列實驗數據資料…………………………………………38
5-2 第一型交互雜交表現量的相關分析…………………………39
5-3隨機抽選的探針組………………………………………43
5-4第二型交互雜交表現量的相關分析……………………44
5-5 本章總結...........................................47
第六章 結論及未來發展……………………………………………49
6-1 研究結論………………………………………………………49
6-2 研究貢獻...........................................50
6-3 建議及後續研究方向…………………………………………52
參考文獻………………………………………………………………55
附錄一..................................................64
圖 目 錄
圖 1.1 研究流程圖……………………………………………………7
圖 2.1 cDNA 微陣列上一個探針的序列………………………………9
圖 2.2 cDNA 微陣列實驗過程………………………………………11
圖 3.1 建構探針基因對應資料庫流程………………………………18
圖3.2 BLAST 比對畫面………………………………………………20
圖 3.3 BLAST比對結果………………………………………………21
圖 3.4 相似性序列資訊………………………………………………22
圖 3.5 UniGene database連結………………………………………23
圖 3.6擷取相似序列的基因資訊……………………………………24
圖 4.1探針長度分布圖………………………………………………26
圖 4.2探針GC content分布圖………………………………………27
圖 4.3探針與相似基因個數分佈圖…………………………………29
圖 4.4探針PT長度的分布情形………………………………………31
圖 4.5 Unique 型探針的長度分佈…………………………………32
圖 4.6 一個探針對應到兩個基因之PT 長度分布圖………………33
圖 4.7 一個探針對應到兩個基因之OT 長度分布圖………………34
圖 5.1相似長度與相關係數分佈圖…………………………………40
圖 5.2相似長度與相關係數分佈圖(第一型)………………………42
圖5.3隨機抽選的相關係數分佈圖…………………………………44
圖5.4相似長度與相關係數分佈圖(第二型 >500bp)…………46
圖5.5相似長度與相關係數分佈圖(第二型 300~500bp)…………47


表 目 錄
表4.1 探針對應相似基因的個數分佈………………………………28
表4.2 所有探針PT 長度、相似度的分布情形……………………30
表4.3 Unique 型探針的長度及相似度分佈情形……………………32
表4.4 一個探針對應三個相似基因長度與相似度的個數…………35
表4.5 可能發生交互雜交的探針組…………………………………37
表5.1 第一型交互雜交的探針組……………………………………39
表5.2 交互雜交探針組(三個相似基因)…………………………41
表5.3 交互雜交探針組(四個相似基因)…………………………41
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