【摘要】：為深入研究環(huán)形泰勒蟲(Theileria annulata)spm-N基因的功能,以建立一種方便快速檢測環(huán)形泰勒蟲病的診斷方法,本論文對環(huán)形泰勒蟲spm-N基因進行了擴增,并將目的基因與原核表達載體pET30a(+)和真核表達載體PGBKT7進行連接,首先用原核表達質粒轉化BL21(DE3)感受態(tài)細胞,經IPTG誘導表達重組蛋白,獲得了以分泌型表達的重組蛋白,以此作為抗原,建立了診斷環(huán)形泰勒蟲病的間接ELISA診斷方法,該方法與中華泰勒蟲(T.sinensis),瑟氏泰勒蟲(T.sergenti),牛巴貝斯蟲(Babesia bovis)等梨形蟲的陽性血清均不發(fā)生交叉反應,表明具有良好的敏感性和特異性,可以用于環(huán)形泰勒蟲病的診斷和流行病學調查;然后又用真核表達質粒轉化酵母Y187感受態(tài)細胞,檢測其在酵母菌株中的自激活性、對酵母生長的影響及其在酵母細胞中的表達情況。結果顯示,重組質粒在酵母細胞內成功表達,且對酵母細胞的生長沒有影響,無毒害作用,沒有自激活活性,可被用于篩選與環(huán)形泰勒蟲spm-N蛋白相互作用的物質。
[Abstract]:In order to study the function of (Theileria annulata) spm-N gene and to establish a convenient and rapid diagnostic method for the detection of Thaleria circularis, the spm-N gene was amplified in this paper, and the results were as follows: (1) in this paper, the spm-N gene of Taylor ringworm was amplified by PCR. The target gene was ligated with prokaryotic expression vector pET30a () and eukaryotic expression vector PGBKT7. First, the prokaryotic expression plasmid was transformed into BL21 (DE3) competent cells. The recombinant protein was induced by IPTG to express the recombinant protein, and the recombinant protein expressed in secretory form was obtained. Based on this antigen, an indirect ELISA diagnostic method was established for the diagnosis of circular Taylor disease. This method was associated with T.sinensis, T.sergenti, and Tylutella sinensi, and was used as antigen for the diagnosis of Thalerellosis annularis. None of the positive sera of Bovine Babesia (Babesia bovis) and other piriform worms showed cross-reaction, which indicated that they had good sensitivity and specificity, and could be used in the diagnosis and epidemiological investigation of Thalerellosis circialis. Then Eukaryotic expression plasmid was used to transform yeast Y187 competent cells to detect its self-stimulating activity in yeast strain, its effect on yeast growth and its expression in yeast cells. The results showed that the recombinant plasmid was successfully expressed in yeast cells and had no effect on the growth of yeast cells, no toxic effect and no self-activating activity. The recombinant plasmid could be used to screen the substances interacting with the spm-N protein of Taylor ringworm.
【學位授予單位】：內蒙古農業(yè)大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：S852.7

【參考文獻】

相關期刊論文 前10條

1 陳立偉;;奶牛環(huán)形泰勒蟲病的臨床癥狀、病理變化及防治[J];現(xiàn)代畜牧科技;2016年06期

2 楊志強;;牛環(huán)形泰勒蟲病的病理剖檢與實驗室檢驗[J];養(yǎng)殖技術顧問;2014年01期

3 武文君;張映;方禮祿;張鳳;;ELISA技術在畜牧業(yè)中的應用進展[J];飼料博覽;2012年05期

4 簡子健;馬素貞;黃家雨;孫其吉吉;沈炯玉;苗中秋;呂偉;;牛環(huán)形泰勒蟲GST-Tams1融合蛋白間接ELISA檢測方法的建立[J];新疆農業(yè)科學;2011年03期

5 羅晟遠;;牛泰勒蟲病的防治[J];農村科技;2010年06期

6 劉愛紅;關貴全;劉軍龍;李有全;馬米玲;牛慶麗;任巧云;殷宏;羅建勛;;牛泰勒蟲18S rRNA基因序列比較研究[J];畜牧獸醫(yī)學報;2009年07期

7 曹建新;;奶牛焦蟲病的診斷與防治[J];畜禽業(yè);2007年11期

8 費永亮;;牛環(huán)形泰勒氏焦蟲病的診治與預防[J];中國獸醫(yī)寄生蟲病;2007年02期

9 羅建勛,殷宏,關貴全,馬米玲,孫彩琴,魯炳義,白啟,呂文順,陳溥言;小亞璃眼蜱對牛巴貝斯蟲未定種和環(huán)形泰勒蟲傳播的試驗研究[J];畜牧獸醫(yī)學報;2005年11期

10 黨志勝,羅建勛,殷宏,蔣錫仕;環(huán)形泰勒蟲病診斷技術的發(fā)展概況[J];青海畜牧獸醫(yī)雜志;2004年01期

相關碩士學位論文 前2條

1 李凱;環(huán)形泰勒蟲TaSP重組蛋白間接ELISA診斷方法的建立[D];河南農業(yè)大學;2015年

2 王振寶;小亞璃眼蜱的生活史和抗菌多肽活性的研究及牛環(huán)形泰勒蟲病二溫式PCR檢測方法的建立[D];新疆農業(yè)大學;2009年

，

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