本文選題：豬增生性腸炎 + 胞內(nèi)勞森氏菌��； 參考：《湖南農(nóng)業(yè)大學(xué)》2016年碩士論文

【摘要】：豬增生性腸炎是由胞內(nèi)勞森菌引起的一種接觸性傳染性疾病,以腸道增生為主要特征,其臨床癥狀表現(xiàn)為腹瀉、日增重量下降、生長(zhǎng)緩慢等,病理變化主要為腸道出血、腸壁增粗、腸粘膜增厚,隱窩上皮細(xì)胞增生。該病傳播范圍廣,發(fā)病率高,呈世界流行性趨勢(shì),嚴(yán)重影響豬場(chǎng)經(jīng)濟(jì)效益。本研究旨在建立一套能在臨床上高效、準(zhǔn)確、快速檢測(cè)豬增生性腸炎的診斷方法；并實(shí)現(xiàn)對(duì)勞森氏菌基因L10065的克隆與表達(dá)。1、采集湖南省規(guī)�；i場(chǎng)腹瀉豬的糞便進(jìn)行處理,對(duì)其進(jìn)行DNA抽提、目的片段巢式PCR擴(kuò)增及擴(kuò)增條件的優(yōu)化,然后通過(guò)敏感性、特異性、重復(fù)性實(shí)驗(yàn)及臨床樣本的檢測(cè)驗(yàn)證此方法的可行性。對(duì)巢式PCR檢測(cè)為陽(yáng)性的豬進(jìn)行剖檢,取其小腸做成石蠟切片,進(jìn)行HE染色與改良銀染,分析其組織病理學(xué)變化。結(jié)果成功擴(kuò)增出大小為263bp目的片段,與勞森菌全序列比對(duì)同源性高達(dá)99%。臨床檢測(cè)陽(yáng)性率達(dá)25%,檢測(cè)穩(wěn)定可靠、重復(fù)性好、特異性高；HE染色發(fā)現(xiàn)小腸絨毛內(nèi)杯狀細(xì)胞明顯減少甚至消失,改良銀染發(fā)現(xiàn)小腸隱窩的非正常融合,且隱窩上皮細(xì)胞胞漿內(nèi)可見(jiàn)疑似長(zhǎng)桿狀或彎曲桿狀胞內(nèi)勞森氏菌的扎堆生長(zhǎng)。2、本實(shí)驗(yàn)擴(kuò)增的目的片段L10065全長(zhǎng)750bp,共編碼249個(gè)氨基酸,編碼蛋白分子量為29.2KD。生物信息學(xué)分析發(fā)現(xiàn),L10065編碼的蛋白具有不穩(wěn)定性,為非信號(hào)肽,預(yù)測(cè)可能是參與細(xì)胞信號(hào)傳導(dǎo)的膜受體蛋白。SDS-PAGE凝膠電泳鑒定L10065編碼的蛋白為可溶性蛋白。
[Abstract]:Porcine proliferative enteritis is a kind of contact infectious disease caused by intracellular Lawson's bacteria. It is characterized by intestinal hyperplasia. Its clinical symptoms are diarrhea, daily weight loss, slow growth and so on. The pathological changes are mainly intestinal bleeding. Thickening of intestinal wall, thickening of intestinal mucosa and proliferation of crypt epithelial cells. The disease has a wide spread range and a high incidence, which has a worldwide epidemic trend and seriously affects the economic benefits of pig farms. The purpose of this study was to establish a highly effective, accurate and rapid diagnostic method for detecting porcine proliferative enteritis, and to clone and express L10065 gene of Lawson's bacillus, and to collect feces from diarrhea pigs in large-scale pig farms in Hunan Province. The DNA was extracted and the nested PCR amplification conditions were optimized. The feasibility of the method was verified by sensitivity, specificity, reproducibility and clinical sample detection. The piglets with positive nested PCR were dissected, and their small intestine was made into paraffin sections. The histopathological changes were analyzed by HE staining and improved silver staining. Results the target fragment of 263bp was amplified successfully, and the homology of the fragment was as high as 99g with the whole sequence of Lawson's bacillus. The positive rate of clinical examination was 25%, the detection was stable and reliable, reproducible, specific and HE staining showed that goblet cells in small intestinal villi decreased or even disappeared, improved silver staining found abnormal fusion of intestinal crypt. In addition, in the cytoplasm of crypt epithelial cells, the growth of L10065 is 750bp, encoding 249 amino acids, and the molecular weight of encoded protein is 29.2kD. the growth rate of L10065 is 750bp. the length of L10065 is 750bp, and the molecular weight of protein is 29.2kD. Bioinformatics analysis showed that the protein encoded by L10065 was unstable and non-signal peptide. The membrane receptor protein, SDS-PAGE, which might be involved in cell signal transduction, was identified as soluble protein by SDS-PAGE gel electrophoresis.
【學(xué)位授予單位】：湖南農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類號(hào)】：S858.28

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本文編號(hào)：1962571