Analysis of the Human Oral Microbiome of Smokers and Non-Smokers Using PCR-RFLP and Ribotyping

Recent advances in the field of microbial and medical ecology emphasize the critical role played by oral bacteria in the delicate dynamic equilibrium of human health and disease, creating the need to define the bacterial communities associated with healthy and non-healthy conditions and to capture shifts in community structure germane to diagnosis. Employing PCR-RFLP of the 16S rDNA gene from metagenomes and plate-wash (cultured) bacteria of oral wash from 10 volunteers, this study evaluated the stability of oral bacteria in healthy subjects and documented community shifts in smokers. Sequence analysis of selected 16S gene amplicons cloned with the Gene Hunter PCR-Trap vector and pCR 4-TOPO cloning kits was conducted to determine the bacteria identity and diversity indices of the two groups. Ribopatterns generated by the restriction enzymes HaeIII and Sau3AI were significantly (p AluI using the GelCompare II software cluster analysis. A stable core of bacteria DNA fingerprint was detected in all healthy subjects, and remained unchanged over the study period of 3 months. Signature bands (1500 bp with HaeIII) in smokers and in non-smokers (800 bp and 700 bp with Sau3A1) were evidently suggesting the presence of potential biomarkers of healthy and non-healthy states. There was no significant difference in the DNA fingerprints of cultured and metagenomic extracts. The genera Xanthomonas, Streptococcus and phylum Candidatus occurred in large numbers in both groups, however, a major shift in composition with the dominance of gram-negative bacteria in smokers compared to healthy subjects was quite remarkable. Taxonomic diversity in smokers was quite high, including members of the genera Rothia, Synechococcus, Neisseria, Thiomargarita and Pyrobaculum. These data highlight the presence of a stable core microbiome amidst a wide diversity, identify a distinct smokers’ cluster and open the way for the search for potential biomarkers for specific diseases.

生物信息流中序列分析软件的设计与开发

介绍了基于Java语言的跨平台生物信息分析平台的设计与实现过程,并对软件系统结构、功能模块、关键技术进行了阐述。该系统能完成生物信息流中序列分析工作。它的开发为从事生物信息学研究的人员提供了有效的数据处理和分析工具。

限制性内切酶分析法在柯萨奇B组病毒检测及分型中的应用

目的 探讨限制性内切酶分析法在柯萨奇B组病毒B1～B6型检测及分型中的应用。方法 利用GCG软件对柯萨奇B1～B6型病毒cDNA全序列进行了限制性内切酶酶切位点的分析。结果 共 114种限制性内切酶在柯萨奇B1～B6型病毒cDNA中分布有不同数量的酶切位点。经过比较和分析 ,得到了它们的一个共同cDNA片段 ,在这个cDNA片段中 ,B1～B6型病毒各自具有特异性的核苷酸序列 (特异的限制性内切酶位点 ) ,对柯萨奇B组病毒 6个已知型别标准毒株进行了成功的分型和检测。结论 本方法有效地解决了在柯萨奇B组病毒检定以往方法中所存在的非特异性问题 ,可以对柯萨奇B组病毒进行准确的检定和型别鉴定。

基于沙眼衣原体ompl基因限制性片段长度多态性和测序法用于女性生殖道感染沙眼衣原体基因分型

沙眼衣原体D-K型见于女性生殖道感染,尤其是宫颈感染的主要病原体,基因分型针对编码主外膜蛋白(MOMP)编码基因(ompl)多态性,CT各型ompl基因长度略有差异,总长度均在1.1 kb左右,使用ompl基因限制性片段长度多态性(restriction fragment length polymorphism,RFLP)进行分型,但是不同的引物设计和实验条件,结果的判读不同,结合ompl基因测序法,能构建出不同实验条件下PCR-RFLP酶切图谱,便于临床判读,并且通过BLASTA及多序列比对发现ompl基因序列碱基变异。收集2010年1月至2014年5月在柳州市人民医院就诊的宫颈脱落细胞沙眼衣原体阳性女性,结合omp1基因测序法进行分型,建立反应体系,通过ompl基因测序结果比对,确定本实验条件的参考酶切图谱,用于临床标本的CT分型检测。基于ompl基因的PCR-RFLP技术是目前沙眼衣原体基因分型临床可运用的方法。

乙型肝炎病毒核心基因的酶切图谱及核苷酸序列分析

对慢性乙型肝炎患者的２２份肝穿刺活检组织及其中配对的６份血清用ＰＣＲ扩增出乙型肝炎病毒（ＨＢＶ）的ＰｒｅＣ／Ｃ基因，选用ＡｖａⅡ、Ｓａｕ３ＡⅠ、ＸｍｎＩ，ＢｓｔＮＩ及ＴａｑＩ５种限制性内切酶消化后，对Ｃ基因进行酶谱分析。发现有６份肝组织及１份血清出现异常酶谱。对Ｓａｕ３ＡⅠ酶解异常标本进行分子克隆及核苷酸序列分析，发现２例患者因２１３７位核苷酸点突变出现了Ｓａｕ３ＡⅠ的新酶切点。类似变化在肝癌组织的ＨＢＶＣ基因中也曾发现，其意义待进一步研究。