本實驗室以研究吳郭魚之神經發育為主軸，先前的研究已建立了吳郭魚腦的
表現序列標籤資料庫(expressed sequence tags, ESTs)，從中取得具神經發育相關性
基因High mobility group proteins 2 (HMG2)，並對其進行生物資訊學分析以及其相
關功能性之探討。HMG2 是屬於不具組蛋白的染色質蛋白質(non-histone chromatin
protein)。本研究利用生物資訊學的方法，針對HMG2 進行開放閱讀區(open reading
frame, ORF)分析，得到642bps 的DNA 序列，並轉譯出213 個胺基酸序列，使用
NCBI Conserved Domain search service 分析其DNA 結合位以及保守區域，並利用
NEBcutter V2.0 對其進行限制酵素的切位分析。另，將HMG2 的ORF 序列於NCBI
與其他物種進行比對，並用胺基酸序列繪製親緣關係樹狀圖(Neighbor Phylogenetic
Tree)，藉以瞭解與其他物種的親緣關係。另一方面，利用聚合酶連鎖反應(PCR)
之方法，建構HMG2 之ORF、建構 pRSET-HMG2 質體、以原核生物系統 BL21
E.coli 來表現HMG2 重組蛋白質，並藉由以6X His Tag 標定的親和性管柱層析方
法(affinity chromatography)進行蛋白質純化，以作為後續 HMG2 蛋白質相關研究
之應用。

The expressed sequence tags, derived from the developing tilapia brain, were established in our laboratory. An unique gene, high mobility group proteins 2 (HMG2), were investigated. HMG2 is a non-histone chromatin protein. HMG2 cloned from tilapia, Oreochromis mossambicus, is a gene with open reading frame 642bps, and deduced as 213 amino acid protein sequence. NCBI Conserved Domain search, the Gene Ontology, NEBcutter restrict enzyme analysis, NCBI blastx, and neighbor phylogenetic tree were used for the bioinformatic analysis. In the present study, the protein expression of HMG2 in the BL21 E.coli system, a prokaryotic system, and purified with 6X His Tag NI-NTA affinity chromatography.

中文摘要....................................................................................................ii
英文摘要...................................................................................................iii
目錄……………………………………………………………………...iv
附圖、表之目錄…………………………………………………………v

前言………………………………………………………………………1
實驗材料與方法………………………………………………………....4
結果……………………………………………………………………..17
討論……………………………………………………………………..20
參考文獻………………………………………………………………..23
附圖、表………………………………………………………………..28
附錄……………………………………………………………………..43

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