空肠弯曲菌铁相关受体基因序列的分析研究

发布时间：2018-07-08 11:46

本文选题：空肠弯曲菌 + 铁相关受体　；参考：《河北医科大学》2011年硕士论文

【摘要】：目的:通过比对空肠弯曲菌(Campylobacter jejuni, C.jejuni)的CfrA,ferric enterobactin receptor,铁吸收相关受体的基因序列,分析CfrA序列核酸及氨基酸序列特征,了解CfrA基因的突变碱基,蛋白质二级结构,疏水性特点,推测可能存在的高变异区段,同时分析其遗传进化关系,为C.jejuni疫苗研究提供基础资料。方法:选取分离自河北医科大学附属第二医院神经内科临床确诊感染C.jejuni的患者粪便并且已经由动物模型证实为致吉兰-巴雷综合征的3株C.jejuni它们分别为zhanxing strain,, lulei strain, qiaoyuntao strain,以及选取来自NCBI的GeneBank中12株的已经测定CfrA的空场弯曲菌CfrA的基因序列。将本地三株C.jejuni菌株进行细菌复苏,培养,提取基因组DNA并测序。应用SequenceAlignment生物信息学软件,将得到的全基因组序列与Genbank中NCTC11168序列对照进行比对,了解CfrA基因的突变碱基。应用DNAstar Protean软件对其编码的蛋白质结构进行对比分析,了解CfrA基因的蛋白质二级结构、疏水性特点。应用MegAlign软件,计算序列间的遗传距离。将得到的12株C.jejuni菌株的CfrA基因序列根据遗传树分组,应用DNASTAR Editseq,MegAlign等生物信息学软件对CfrA基因序列进行组间的比较,推测可能存在的高变异区段。应用DNAstar Protean软件对其编码的蛋白质结构进行对比分析,了解CfrA二级结构、疏水性特点。并分析其遗传进化关系. 结果:本地zhanxing strain,lulei strain,qiaoyuntao strain的CfrA基因均由2091个碱基构成,与NCTC11168的cfra基因序列相比,突变率分别为0%;1.4%; 1.3%。疏水性无明显变化。遗传距离计算最大为zhanxing株与lulei株距离为1.6%。来自NCBI十二株空肠弯曲杆菌cfra基因中(1)C. jejuni subsp. jejuni 84-25 gcj8425_13(2)C.jejuni subsp. jejuni 260.94c_jejuni_subsp_jejuni_26094_40(3)C.jejuni subsp. doylei 269.97(4) C.jejuni subsp. jejuni CG8421 C493四株存在插入与缺失。余序列均由2091个碱基构成,最高变异率为6.79%.碱基高变异的区段分别为,第一组有高度相似性,高变异区段为6-147,第二组变异率较高,变异区段1-1008保守区段在1059-2075,第三组高变异;蛋白质二级结构,第1组5-54位,524-692位区段二级结构有变异。第2组1-24位区段520-692位区段二级结构有变异。各个区段位于表面的可能性均较小。疏水性均无明显变化。遗传距离计算最大为S3B株与M76297株距离为5.5%。结论:15株菌株除四株外均显示较高的保守性。对它们的序列同源性进行了对比,表明Cfra基因的同一性的水平波动在93%-100%。zhanxing strain, lulei strain, qiaoyuntao strain遗传距离较小,可能反映了河北地区空肠弯曲菌在进化上存在一定的聚类现象,推测具有的区域特征。第三组菌株的高变异情况,推测空肠弯曲菌铁摄取系统的表面受体基因可能不是单一的,而是多个基因共同作用的结果;或者,菌株可能含有同源性铁调节外膜蛋白,总之本研究中的15株菌株的序列对比的情况为建立研究华北地区菌株的地域性特点及对空肠弯曲菌感染的监测及疫苗的制备奠定了基础。
[Abstract]:Objective: to analyze the nucleotide and amino acid sequence characteristics of CfrA sequence by comparing the CfrA, ferric enterobactin receptor, and iron absorption related receptor of Campylobacter jejuni (C.jejuni), and to understand the mutation base of the CfrA gene, the two stage structure of the protein and the hydrophobicity of the protein, and to speculate on the possible high variation section. The genetic evolution relationship was analyzed to provide basic information for C.jejuni vaccine research.
Methods: the feces of patients who were isolated from the neurology department of the Second Affiliated Hospital of Hebei Medical University were selected and 3 strains of C.jejuni, which were confirmed to be Gillain Barre syndrome, were Zhanxing strain, Lulei strain, qiaoyuntao strain, and 12 strains of GeneBank from NCBI were selected. The gene sequence of CfrA of Campylobacter CfrA was measured. Three local strains of C.jejuni strains were resuscitation, cultured, and genomic DNA was extracted and sequenced. The whole genome sequence was compared with the NCTC11168 sequence in Genbank by SequenceAlignment bioinformatics software, and the mutation base of the CfrA gene was understood. DNAst AR Protean software contrasts its encoded protein structure to understand the two level structure and hydrophobicity of the protein of the CfrA gene, and uses the MegAlign software to calculate the genetic distance between the sequences.
The CfrA gene sequences of 12 strains of C.jejuni strain were grouped according to the genetic tree, and the DNASTAR Editseq, MegAlign and other bioinformatics software were used to compare the sequences of CfrA genes to speculate on the possible high variation section. The DNAstar Protean software was used to compare and analyze the egg white structure encoded by the DNAstar Protean software, and to understand the CfrA two junction. The characteristics of hydrophobicity were analyzed and their genetic evolution relationship was analyzed.
Results: the CfrA gene of local Zhanxing strain, Lulei strain and qiaoyuntao strain was composed of 2091 bases. Compared with the CFRA gene sequence of NCTC11168, the mutation rate was 0% and 1.4%, and the hydrophobicity of 1.3%. was not obviously changed. The maximum distance between Zhanxing strain and Lulei strain was from Twelve Strains of Campylobacter jejuni. The gene (1) C. jejuni subsp. jejuni 84-25 gcj8425_13 (2) C.jejuni subsp. jejuni 260.94c_jejuni_subsp_jejuni_26094_40 (3) C.jejuni subsp. doylei 269.97 (4) exists insertion and deletion. The residual sequence is composed of 2091 bases, and the highest variation rate is the section of high variation of base base. The first group is highly similar, the high variation section is 6-147, the second groups have a high variation rate, the 1-1008 conservative section of the variation section is 1059-2075, and the third groups are high variation; the protein two structure, first group 5-54, and 524-692 bit section two structure have variation. The second groups of 520-692 section of 520-692 districts have variation. Each section is located in the section. There was no significant change in the surface hydrophobicity. The largest genetic distance was S3B and M76297 5.5%..
Conclusion: the 15 strains of the 15 strains showed high conservatism, and their sequence homology showed that the genetic distance of the homogeneity of the Cfra gene was fluctuating in 93%-100%.zhanxing strain, Lulei strain and qiaoyuntao strain, which may reflect the evolution of Campylobacter jejuni in Hebei. The high variation of the third groups of strains, speculates that the surface receptor gene of the ferric uptake system of Campylobacter jejuni may not be single, but the result of multiple genes, or the strain may contain homologous iron regulating outer membrane protein, and in a word, the sequence comparison of the 15 strains in this study The study laid a foundation for establishing the regional characteristics of the strains in North China and monitoring the Campylobacter jejuni infection and the preparation of the vaccine.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R378

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