基于液态悬浮芯片的鼠疫耶尔森氏菌SNP分型
发布时间：2018-11-17 06:44
【摘要】：鼠疫耶尔森氏菌(Yersinina pestis,以下简称鼠疫菌)曾引发世界范围的三次鼠疫大爆发,给人类带来了灾难。鼠疫之所以能够如此猖獗的肆掠整个世界,主要是因为鼠疫作为自然疫源性疾病以其传播速率极快,突发病急不可预测,死亡率极高和传染性极强为特征的甲类烈性传染病。本研究利用多种遗传标记物对我国鼠疫东方型菌株进行分型,可以避免采用单一遗传标记而导致的分辨率不足或偏性结果。主要研究结果如下： 1.建立了Luminex多重检测SNP的技术平台。鼠疫菌作为一种较为“年轻”且缺少基因重组的病原细菌,单核苷酸多态性(Single Nucleotide polymorphisms, SNP)可作为其非常可靠分型和系统发育分析指标。本研究建立基于Lumienx x-TAG微球(液态悬浮芯片)的鼠疫菌SNP分型技术,利用多重PCR同时扩增鼠疫菌18个SNP位点片段,将扩增完成的产物进行多重ASPE (Allele Specific Primer Extension)反应以及Luminex技术多重SNP检测,通过一次反应可以检测出待测菌株中18个SNP位点的碱基特征。利用这一方法分析20株鼠疫东方型菌株,可以将其根据18个SNP位点的特征分为7个基因型,本方法具有判读方便快速、高通量、成本低廉等特点,在病原菌的SNP分型方面具有广阔的应用前景。 2.初步探讨了我国鼠疫菌东方型的起源。目前国际上普遍认为,鼠疫的第三次大流行起源于中国云南,鼠疫菌东方型菌株也起源于云南。但这与我们前期所研究的结果在某些方面与这一轮点不相符,即我们目前所测定的云南东方型菌株都是比较“年轻”的菌株,而中国最古老的东方型菌株分离自福建。本研究将运用Luminex技术平台检测分析大量的云南东方型菌株,对云南东方型菌株有个初步的探讨分型结果,我们对193株鼠疫东方型菌株进行分型,一共分为13个基因型,分别是a, b, c, d, e, f, g, h, i, j, k,l, m。53个SNP位点信息对后期期追溯东方型鼠疫菌的起源提供了重要的数据。
[Abstract]:Yersinia pestis (Yersinina pestis, (Yersinia pestis) has caused three outbreaks of Yersinia pestis worldwide and brought disaster to human beings. The reason why plague is rampant throughout the world is mainly because plague as a natural epidemic disease is characterized by its rapid transmission rate, sudden and unpredictable disease, very high mortality and highly infectious type A strong infectious disease. In this study, a variety of genetic markers were used to type the oriental strains of plague in China, which could avoid the lack of resolution or bias caused by the use of single genetic marker. The main results are as follows: 1. The technical platform of Luminex multiple detection SNP is established. Yersinia pestis as a relatively "young" and lack of gene recombination of pathogenic bacteria, single nucleotide polymorphism (Single Nucleotide polymorphisms, SNP) can be used as a very reliable typing and phylogenetic analysis index. In this study, the SNP typing technique of Yersinia pestis based on Lumienx x-TAG microspheres (liquid suspension chip) was established, and 18 SNP loci of Yersinia pestis were amplified simultaneously by multiple PCR. The amplified products were detected by multiple ASPE (Allele Specific Primer Extension) reaction and Luminex technique by multiple SNP detection. The base characteristics of 18 SNP sites in the tested strain could be detected by one reaction. By using this method, 20 strains of Yersinia pestis oriental strain can be divided into 7 genotypes according to the characteristics of 18 SNP loci. The method is characterized by easy and fast interpretation, high throughput and low cost. It has a broad application prospect in SNP typing of pathogenic bacteria. 2. The oriental origin of Yersinia pestis in China was preliminarily discussed. At present, it is widely believed that the third pandemic of plague originated in Yunnan, China, and the oriental strain of Yersinia pestis also originated in Yunnan. However, this does not accord with the results of our previous studies in some respects, that is, the oriental strains in Yunnan are relatively "young" strains, and the oldest oriental strains in China are isolated from Fujian Province. In this study, a large number of oriental strains of Yunnan Province were detected and analyzed by using Luminex technology platform, and a preliminary study on the results of typing of oriental strains of Yunnan Province was carried out. We typed 193 strains of Oriental Yersinia pestis into 13 genotypes. The information of SNP loci of a, b, c, d, e, f, g, h, i, j, kanll and m.53 provides important data for tracing the oriental Yersinia pestis origin in late stage.
【学位授予单位】：湖南农业大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R378
本文编号：2336811
[Abstract]:Yersinia pestis (Yersinina pestis, (Yersinia pestis) has caused three outbreaks of Yersinia pestis worldwide and brought disaster to human beings. The reason why plague is rampant throughout the world is mainly because plague as a natural epidemic disease is characterized by its rapid transmission rate, sudden and unpredictable disease, very high mortality and highly infectious type A strong infectious disease. In this study, a variety of genetic markers were used to type the oriental strains of plague in China, which could avoid the lack of resolution or bias caused by the use of single genetic marker. The main results are as follows: 1. The technical platform of Luminex multiple detection SNP is established. Yersinia pestis as a relatively "young" and lack of gene recombination of pathogenic bacteria, single nucleotide polymorphism (Single Nucleotide polymorphisms, SNP) can be used as a very reliable typing and phylogenetic analysis index. In this study, the SNP typing technique of Yersinia pestis based on Lumienx x-TAG microspheres (liquid suspension chip) was established, and 18 SNP loci of Yersinia pestis were amplified simultaneously by multiple PCR. The amplified products were detected by multiple ASPE (Allele Specific Primer Extension) reaction and Luminex technique by multiple SNP detection. The base characteristics of 18 SNP sites in the tested strain could be detected by one reaction. By using this method, 20 strains of Yersinia pestis oriental strain can be divided into 7 genotypes according to the characteristics of 18 SNP loci. The method is characterized by easy and fast interpretation, high throughput and low cost. It has a broad application prospect in SNP typing of pathogenic bacteria. 2. The oriental origin of Yersinia pestis in China was preliminarily discussed. At present, it is widely believed that the third pandemic of plague originated in Yunnan, China, and the oriental strain of Yersinia pestis also originated in Yunnan. However, this does not accord with the results of our previous studies in some respects, that is, the oriental strains in Yunnan are relatively "young" strains, and the oldest oriental strains in China are isolated from Fujian Province. In this study, a large number of oriental strains of Yunnan Province were detected and analyzed by using Luminex technology platform, and a preliminary study on the results of typing of oriental strains of Yunnan Province was carried out. We typed 193 strains of Oriental Yersinia pestis into 13 genotypes. The information of SNP loci of a, b, c, d, e, f, g, h, i, j, kanll and m.53 provides important data for tracing the oriental Yersinia pestis origin in late stage.
【学位授予单位】：湖南农业大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R378
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