本文選題：布魯氏菌 + BMEII基因��； 參考：《生物技術(shù)》2017年02期

【摘要】：[目的]檢測布魯氏菌BMEII0988基因的原核表達(dá)情況并對其免疫原性進(jìn)行分析。[方法]以熱滅活的布魯氏菌16M標(biāo)準(zhǔn)株為模板,根據(jù)Gen Bank中公布的羊種布魯氏菌16M的BMEII0988基因序列分別設(shè)計引物,用PCR方法擴(kuò)增BMEII0988基因的編碼序列,連接到T載體測序,將測序正確的基因序列克隆到原核表達(dá)載體p ET-32a上,轉(zhuǎn)化入大腸桿菌DE3感受態(tài)細(xì)胞中誘導(dǎo)表達(dá),將獲得的目標(biāo)蛋白用AKTAxpress智能多維純化系統(tǒng)進(jìn)行純化,并用Western Blot分析其反應(yīng)原性。[結(jié)果]BMEII0988基因長度為1 131 bp,編碼377個氨基酸,SDS-PAGE表明BMEII 0988融合蛋白在大約66 k Da處出現(xiàn)條帶,純化后條帶單一,BMEII 0988蛋白(NCBI標(biāo)準(zhǔn)序列號:WP_002966326.1)具有較好的反應(yīng)原性。[結(jié)論]該研究為下一步建立相應(yīng)蛋白標(biāo)記的診斷方法和疫苗的研發(fā)奠定了基礎(chǔ)。
[Abstract]:[objective] to detect the prokaryotic expression of brucella BMEII0988 gene and analyze its immunogenicity. [methods] the 16M standard strain of brucella was used as template, primers were designed according to the 16M BMEII0988 gene sequence published in Gen Bank, the coding sequence of BMEII0988 gene was amplified by PCR method, and ligated to T vector sequencing. The correctly sequenced gene sequence was cloned into prokaryotic expression vector p ET-32a and transformed into Escherichia coli DE3 receptive cells to induce expression. The target protein was purified by AKTAxpress intelligent multidimensional purification system and its reactivity was analyzed by Western Blot. [results] the length of BMEII0988 gene was 1 131 BP, encoding 377 amino acids. SDS-PAGE showed that the fusion protein of BMEII 0988 appeared bands at about 66kDa. After purification, the single sequence number of BMEII 0988 protein was: WP002966326.1). [conclusion] this study lays a foundation for the next step to establish the diagnostic method of the corresponding protein marker and the development of vaccine.
【作者單位】： 銅仁學(xué)院大健康學(xué)院;銅仁學(xué)院農(nóng)林工程與規(guī)劃學(xué)院(銅仁市文化科技產(chǎn)業(yè)創(chuàng)新研究中心);浙江大學(xué)動物科技學(xué)院;石河子大學(xué)動物科技學(xué)院;商丘師范學(xué)院生物與食品學(xué)院;邢臺市第二醫(yī)院肝病二科;石河子大學(xué)醫(yī)學(xué)院;
【基金】：國家自然科學(xué)基金項目(“E2泛素結(jié)合酶關(guān)鍵分子調(diào)控胞內(nèi)布魯氏菌存活的分子機(jī)制研究”,No.31502067;“布魯氏菌轉(zhuǎn)錄因子GntR調(diào)控細(xì)胞凋亡的分子機(jī)制研究”,No.31602080) 銅仁學(xué)院博士科研啟動基金項目(“布魯氏菌感染巨噬細(xì)胞后PI3K/Akt通路調(diào)控凋亡的分子機(jī)制研究”,No.trxy DH1504) 貴州省教育廳自然科學(xué)研究重點(diǎn)項目(“Toll樣受體信號通路在綿羊肺炎支原體感染宿主細(xì)胞中的作用機(jī)理研究”,黔教合KY字(2015)411號) 貴州省科技合作計劃項目(“TLR2/6-NF-κB信號通路在絲狀支原體山羊亞種感染中的機(jī)制研究”,黔科合LH字[2015]7238);貴州省科技合作計劃項目(“JAK/STAT信號通路在單核細(xì)胞增生李斯特菌侵染腦微血管內(nèi)皮細(xì)胞中的作用機(jī)制研究”,黔科合LH字[2015]7236號)
【分類號】：R378.5;S852.61
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本文編號：1916034