發(fā)布時(shí)間：2019-02-24 17:34

【摘要】：MD是由MDV感染引發(fā)的一種常見(jiàn)的惡性淋巴細(xì)胞瘤傳染病。為進(jìn)一步研究MDV的致瘤機(jī)制,本研究通過(guò)MDV超強(qiáng)毒株GX0101感染SPF雞群構(gòu)建動(dòng)物模型:(1)將1日齡的SPF雛雞分為試驗(yàn)組和對(duì)照組各40只,試驗(yàn)組每只雛雞腹腔注射200μL的GX0101型MDV毒株,對(duì)照組每只雛雞等量注射CEF原代細(xì)胞液體。此后在第7 d、14 d、21 d、30 d、45 d和60 d各采取3只試驗(yàn)組和對(duì)照組雞的心、肝、脾、肺、腎、腺胃、胸腺,腦,胰腺等組織樣品和血液,通過(guò)熒光定量PCR檢測(cè)不同時(shí)間點(diǎn)血液中meq和gB的表達(dá)量,通過(guò)HE染色方法觀察各組織病變情況;(2)通過(guò)Label-free技術(shù)分析了45 d后試驗(yàn)組和對(duì)照組雞脾臟的蛋白質(zhì)組學(xué),并對(duì)差異蛋白進(jìn)行了GO分析和KEGG分析;(3)通過(guò)熒光定量PCR研究了注射GX0101毒株和CEF細(xì)胞后不同時(shí)間點(diǎn)試驗(yàn)組和對(duì)照組雞各組織的PRNP相對(duì)表達(dá)量。通過(guò)本研究得出結(jié)論:(1)熒光定量PCR檢測(cè)發(fā)現(xiàn),MDV標(biāo)志性基因meq和gB在注射GX0101后45 d表達(dá)量最高,并多個(gè)組織在45 d觀察到了腫瘤,meq和gB表達(dá)量與腫瘤的形成相關(guān);(2)試驗(yàn)組同對(duì)照組相比,其中有690個(gè)蛋白表達(dá)上調(diào),390個(gè)蛋白下調(diào)。GO分析注釋結(jié)果顯示,這些差異蛋白涉及到的生物過(guò)程主要是代謝過(guò)程,轉(zhuǎn)運(yùn)和氧化還原過(guò)程。KEGG經(jīng)典通路分析顯示這些差異蛋白主要涉及到癌癥相關(guān)的通路,朊蛋白相關(guān)的通路,信號(hào)通路以及其它通路等。(3)PRNP基因表達(dá)量與MDV感染具有相關(guān)性,并測(cè)得PRNP基因在試驗(yàn)組雞群多個(gè)組織中的表達(dá)量基本高于對(duì)照組雞群。PRNP基因在注射組多個(gè)組織的表達(dá)量隨著時(shí)間的變化而變動(dòng),在45 d達(dá)到峰值,此后逐漸下降至60d并處于較低的轉(zhuǎn)錄水平。
[Abstract]:MD is a common malignant lymphocytoma infection caused by MDV infection. In order to further study the tumorigenic mechanism of MDV, we established an animal model of SPF chickens infected with MDV supervirulent GX0101 strain. (1) the 1-day-old SPF chicks were divided into two groups: the experimental group and the control group (n = 40). In the experimental group, 200 渭 L GX0101 type MDV strain was injected intraperitoneally in each chick, while in the control group, the primary cell fluid of CEF was injected into the same quantity. After that, the heart, liver, spleen, lung, kidney, glandular stomach, thymus, brain, pancreas and other tissue samples and blood of 3 chickens in the experimental group and the control group were taken at the 7th day, 14 days, 21 days, 30 days, 45 days and 60 days, respectively. The expression of meq and gB in blood at different time points were detected by fluorescence quantitative PCR, and the pathological changes of tissues were observed by HE staining. (2) the proteomics of spleen of the experimental group and the control group were analyzed by Label-free technique after 45 days, and the differential proteins were analyzed by GO and KEGG. (3) the relative expression of PRNP in chicken tissues at different time points after injection of GX0101 and CEF cells was studied by fluorescence quantitative PCR. The results of this study were as follows: (1) fluorescence quantitative PCR analysis showed that the expression of MDV signature genes meq and gB was the highest at 45 days after GX0101 injection, and the tumor was observed in several tissues at 45 days. The expression of meq and gB was related to the formation of tumor. (2) compared with the control group, 690 proteins were up-regulated and 390 proteins down-regulated in the experimental group. The results of GO analysis showed that the biological processes involved in these differential proteins were mainly metabolic processes. Transport and redox processes. KEGG classic pathway analysis shows that these differential proteins are mainly involved in cancer-related pathways, prion protein-related pathways, (3) the expression of PRNP gene was correlated with MDV infection. The expression of PRNP gene in multiple tissues of experimental group was higher than that in control group. The expression of PRNP gene in multiple tissues of injection group changed with time and reached its peak at 45 days. After that, it gradually decreased to 60 days and was at a lower level of transcription.
【學(xué)位授予單位】：甘肅農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：S858.31

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