目录
摘要Ⅱ
关键词Ⅱ
AbstractⅡ
引言
引言
1 材料与方法1
1.1前期准备 1
1.2 DNA提取 1
1.3克隆文库构建1
1.3.1 16S rDNA富集 1
1.3.2 连接转化克隆文库2
1.4 限制性内切酶分型3
1.4.1 挑选阳性克隆并进行片段扩增 .3
1.4.2连接转化克隆文库3
1.4.3 电泳检测及带型分析 .4
1.5 系统发育学分析方法3
2 结果与分析5
2.1电泳图带型分析5
2.2 通过RFLP手段鉴定出的细菌种类5
2.3灰飞虱体内微生物系统发育学分析6
2.4灰飞虱体内细菌的相对丰度6
3 讨论.. 11
致谢13
参考文献14
图21 PCR产物酶切分析结果5
图22所有阳性克隆得到的微生物种类组成7
图 23 基于RFLP的灰飞虱体内细菌微生物系统发育树8
图 24 基于RFLP的灰飞虱体内Wolbachia 16S rDNA系统发育树9
图25 基于RFLP的灰飞虱体内Asaia 16S rDNA系统发育树10
表21灰飞虱体内细菌微生物种类组成6
基于RFLP技术的灰飞虱体内细菌微生物多样性的研究
摘要
灰飞虱（Laodelphax striatellus）以刺吸式口器取食植物韧皮部汁液，是重要的水稻害虫。昆虫体内的微生物大多是不可进行体外培养的，因此可以借助限制性内切酶片段长度多态性分型（RFLP）对不同种群的灰飞虱体内细菌微生物群落多样性进行研究。通过研究，我们从灰飞虱体内鉴定出8种细菌，分别是：Acinetobacter soli、Asaia symbiont、Cutibacterium acnes、Klebsiella pneumoniae、Pseudomonas poae、Pseudomonas protegens、Rubellimi *51今日免费论文网|www.51jrft.com +Q: &351916072& 
crobium sp.和Wolbachia symbiont。其中，Wolbachia在灰飞虱体内占主导地位，占所有阳性克隆得到的微生物种类的56.72%。P. poae是第一次在灰飞虱中被检测到。Asaia只在实验室种群体内被检测到，并且在实验室种群体内，Asaia含量高于Wolbachia。我们基于PCRRFLP的方法虽然耗时，但可以克服普通的特异性PCR筛选和高通量测序对灰飞虱体内微生物鉴定的局限性，从而更准确的鉴定灰飞虱体内的共生菌物种。
Research of bacterial and microbial diversity in Laodelphax striatellus based on RFLP technology
Name : Gao Zijie Instructor : Bing Xiaoli
ABSXRACT
Laodelphax striatellus feeds on plant phloem sap with piercingsucking mouthparts and is an important rice pest. Many microorganisms in insects cannot be cultured in vitro. Therefore, the diversity of bacterial communities in different populations of L. striatellus can be studied by using restriction enzyme fragment length polymorphism (RFLP). In this research, we identified 8 kinds of bacteria, namely: Acinetobacter soli, Asaia symbiont, Cutibacterium acnes, Klebsiella pneumoniae, Pseudomonas poae, Pseudomonas protegens, Rubellimicrobium sp. and Wolbachia symbiont. Among them, Wolbachia dominates the body of the Laodelphax striatellus, accounting for 56.72% of all the microorganisms obtained from the positive clones. P. poae was detected for the first time in L. striatellus. Asaia was only detected in the laboratory population, and was higher than Wolbachia in that population. Although our PCRRFLPbased method is timeconsuming, it can overcome the limitations of ordinary specific PCR screening and highthroughput sequencing for the identification of microbes in the planthopper, thereby identify the symbiotic species in the Laodelphax striatellus more accurately.

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