本文選題：陰道毛滴蟲 + 衰老��； 參考：《汕頭大學(xué)》2005年碩士論文

【摘要】：研究目的 (1)克隆原生動(dòng)物陰道毛滴蟲(Trichomonas vaginalis) 長(zhǎng)壽保障基因(Longevity assurance gene 1 LAG1)同源基因:Tv- LAG1。構(gòu)建Tv-LAG1原核表達(dá)重組體及表達(dá)其融合蛋白。(2)在 模式生物陰道毛滴蟲cDNA消減雜交文庫(kù)中,篩選和鑒定陰道毛 滴蟲細(xì)胞周期及衰老相關(guān)基因。材料和方法 (1)從陰道毛滴 蟲cDNA表達(dá)文庫(kù)中篩選LAG1的同源基因,并對(duì)其序列保守結(jié)構(gòu) 域進(jìn)行同源性分析。用pET41表達(dá)載體與Tv-LAG1 cDNA克隆構(gòu)建 原核表達(dá)重組體,在異丙基硫代-β-D-半乳糖苷(isopropylthio- β-D-galactoside IPTG)誘導(dǎo)下表達(dá)目的基因融合蛋白,Ni-NTA親 和層析純化表達(dá)產(chǎn)物,SDS-PAGE鑒定表達(dá)產(chǎn)物的分子量及純化 效果。用純化的融合蛋白免疫家兔,獲得抗血清,采用Western blot 法對(duì)抗體特異性進(jìn)行分析鑒定。(2)采用反轉(zhuǎn)錄PCR技術(shù)進(jìn)一步 分析鑒定cDNA消減雜交文庫(kù)中分離到的15個(gè)cDNA克隆。結(jié)果 (1)從陰道毛滴蟲cDNA表達(dá)文庫(kù)中成功篩選出長(zhǎng)壽保障基因 LAG1的同源基因(Tv-LAG1)。其cDNA克隆含910對(duì)堿基,其編 碼框長(zhǎng)834 bp,推測(cè)蛋白質(zhì)序列含有277個(gè)氨基酸。從植物到人 類的結(jié)構(gòu)域序列分析提示,Tv-Lag1p與酵母Lag1p及其它同源蛋 白均有同源性。其中與擬芥蘭相比,在258個(gè)氨基酸的序列區(qū), 同源性達(dá)到23%。其次為人類Lag1p同源蛋白質(zhì),在216個(gè)氨基酸
[Abstract]:Objective (1) to clone the long assurance gene 1 LAG1 homologous gene of Trichomonas vaginalis. Construction of Tv-LAG1 prokaryotic expression recombinant and expression of its fusion protein. (2) screening and identification of trichomonas vaginalis cell cycle and senescence related genes in the model Trichomonas vaginalis cDNA subtractive hybridization library. Materials and methods (1) the homologous gene of LAG1 was screened from the cDNA expression library of Trichinella vaginalis and its conserved domain was analyzed. The prokaryotic expression recombinant was constructed by using pET41 expression vector and Tv-LAG1 cDNA clone. Isopropylthio- 尾 -D-galactoside (IPTG) was induced by isopropylthio- 尾 -Dgalactoside IPTG to express the target gene fusion protein: Ni-NTA affinity chromatography and SDS-PAGE to identify the molecular weight and purification effect of the expressed product. The antiserum was obtained by immunizing rabbits with the purified fusion protein, and the specificity of the antibody was analyzed by Western blot. (2) 15 cDNA clones isolated from the cDNA subtractive hybridization library were further analyzed by reverse transcription-polymerase chain reaction (RT-PCR). Results (1) the homologous gene (Tv-LAG1) of longevity protection gene LAG1 was successfully screened from the cDNA expression library of Trichomonas vaginalis. The cDNA clone contained 910 pairs of bases, and its coding frame was 834 BP, which suggested that the protein sequence contained 277 amino acids. The analysis of the domain sequence from plant to human indicates that Tv-Lag1p shares homology with yeast Lag1p and other homologous egg whites. The homology of 258 amino acids was 23 in comparison with that of Brassica thaliana. The second is human Lag1p homologous protein, with 216 amino acids.
【學(xué)位授予單位】：汕頭大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2005
【分類號(hào)】：R383

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 章家新,傅玉才,鄭曉虹,劉紅;陰道毛滴蟲cDNA文庫(kù)的構(gòu)建和鑒定[J];汕頭大學(xué)醫(yī)學(xué)院學(xué)報(bào);2003年04期

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本文編號(hào)：2110368