發(fā)布時(shí)間：2018-05-30 02:04

  本文選題：闌尾 + 高通量測(cè)序　； 參考：《南方醫(yī)科大學(xué)》2017年碩士論文

【摘要】：背景與目的人類腸道細(xì)菌有1013-1014個(gè)細(xì)菌,已發(fā)現(xiàn)很多疾病的發(fā)生、發(fā)展都與腸道菌群的異常存在著關(guān)聯(lián),不少學(xué)者開(kāi)始關(guān)注腸道微生物與不同疾病發(fā)病的關(guān)系。闌尾曾被認(rèn)為是在進(jìn)化過(guò)程中殘余的器官,然而,越來(lái)越多的回顧性研究揭示了闌尾切除與腸道疾病發(fā)病的關(guān)系,關(guān)于其機(jī)制的猜想傾向于與闌尾對(duì)腸道微生物和免疫的影響,然而相關(guān)研究有限。目前16SrDNA測(cè)序和宏基因組測(cè)序已廣泛應(yīng)用于微生物的研究。本研究擬用16s rDNA測(cè)序和宏基因組測(cè)序的手段檢測(cè)有闌尾切除手術(shù)史和無(wú)闌尾切除手術(shù)史的受試者的大便標(biāo)本的微生物情況,以研究:有闌尾切除術(shù)史的受試者的大便樣本的微生物物種和功能的結(jié)構(gòu)和豐度,分析其與無(wú)手術(shù)史的受試者的異同,以及探討存在的差異與炎癥性腸病和大腸癌等疾病發(fā)病之間可能的關(guān)系。方法16srDNA擴(kuò)增子測(cè)序分別收集有闌尾切除手術(shù)史(實(shí)驗(yàn)組)和無(wú)闌尾切除手術(shù)史(對(duì)照組)的受試者的大便標(biāo)本1份,分別提取糞便基因組DNA,擴(kuò)增細(xì)菌16srDNAV4片段,采用16srDNA測(cè)序的方法得出測(cè)序信息,經(jīng)GreenGene數(shù)據(jù)庫(kù)確定物種信息,對(duì)細(xì)菌進(jìn)行結(jié)構(gòu)和豐度分析。宏基因組測(cè)序?qū)Σ糠?6srDNA測(cè)序的剩余樣品提取基因組DNA,經(jīng)樣品檢測(cè)、建庫(kù)、宏基因組測(cè)序、數(shù)據(jù)質(zhì)控、組裝、基因預(yù)測(cè)、物種和功能注釋,對(duì)微生物進(jìn)行物種和功能的分析。結(jié)果1.兩組糞便腸道微生物以細(xì)菌為主,占腸道微生物的95%以上,在門(mén)水平均以擬桿菌門(mén),厚壁菌門(mén)和變形菌門(mén)為主;2.兩組糞便微生物組成有微生物群落結(jié)構(gòu)和豐度的差異,其中G_Prevotella,G_Sutterella,S Prevotella bryantii,S_Bacteroides_sp_CAG_530,S_Bacteroides_coprophilu,S_Oscillibacter_sp_ER4,S_Ruminococcus_sp_CAG_177,S_Prevotella_sp_KHD1,S_Clostridium_sp_CAG_138,S_Clostridium_sp_CAG_127,S_Prevotella_sp_CAG_520,S_Bacteroides_plebeius,S_Bacteroides_plebeius_CAG_211 在對(duì)照組中富集,而 S_Bacteroides_fragilis,S_Bacteroides_uniformis在實(shí)驗(yàn)組富集。3.兩組糞便微生物功能有差異,其中闌尾切除術(shù)組中碳水化合物酶富集,與心血管疾病、內(nèi)分泌代謝疾病、次級(jí)代謝物的合成、脂肪代謝通路相關(guān)的基因相對(duì)豐度顯著增加,而與細(xì)胞生長(zhǎng)與死亡、復(fù)制和修復(fù)、翻譯、能量代謝、氨基酸和核酸代謝以及免疫系統(tǒng)等通路相關(guān)的基因相對(duì)豐度明顯降低。結(jié)論闌尾切除改變了腸道微生物的組成和功能,闌尾對(duì)腸道微生物有影響。特殊分類群和基因在闌尾切除術(shù)后組富集,可能與闌尾切除術(shù)后消化系統(tǒng)、心血管系統(tǒng)、內(nèi)分泌系統(tǒng)等疾病有關(guān)。
[Abstract]:Background and objective there are 1013-1014 bacteria in human intestinal tract. Many diseases have been found and the development is related to the abnormality of intestinal flora. Many scholars have begun to pay attention to the relationship between intestinal microbes and the pathogenesis of different diseases. The appendix was once thought to be a residual organ during evolution. However, more and more retrospective studies have revealed the relationship between appendectomy and the pathogenesis of intestinal diseases, and speculation about its mechanism tends to relate to the effects of appendix on intestinal microorganism and immunity. However, the relevant research is limited. At present, 16SrDNA sequencing and macro genome sequencing have been widely used in the study of microbes. In this study, 16 s rDNA sequencing and macro genome sequencing were used to detect the microbial status of stool specimens from subjects with and without appendectomy. To study the structure and abundance of microbial species and function in stool samples of subjects with a history of appendectomy, and analyze their similarities and differences with those without surgical history. And to explore the possible relationship between the differences and inflammatory bowel disease, colorectal cancer and other diseases. Methods one stool specimen with appendectomy history (experimental group) and no appendectomy history (control group) was collected by 16srDNA amplification. Fecal genomic DNA was extracted and bacterial 16srDNAV4 fragments were amplified. The 16srDNA sequencing method was used to obtain the sequencing information. The species information was determined by GreenGene database and the structure and abundance of bacteria were analyzed. Genomic DNA was extracted from the remaining samples of partial 16srDNA sequencing. The species and function of microorganisms were analyzed by sample detection, library construction, macro genome sequencing, data quality control, assembly, gene prediction, species and functional annotation. Result 1. Bacteria were the main microorganism in fecal tract in both groups, accounting for more than 95% of the intestinal microbes, and Bacteroides phylum, Typhylum and Proteus were the main bacteria at the portal level. 涓ょ粍綺�渚垮井鐢熺墿缁勬垚鏈夊井鐢熺墿缇よ惤缁撴瀯鍜屼赴搴︾殑宸�寮，

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