低能离子注入介导的沙漠寡营养细菌rRNA基因突变研究
发布时间：2018-12-20 21:13
【摘要】：低能离子注入生物体技术是近些年来发展起来的物理诱变育种技术,该技术一般采用活性氮离子,注入生物样品后掺入靶分子形成新物质,引起生物体的变异,从而引发生物学效应。为了进一步理解低能N+注入介导外源基因转化沙漠寡营养细菌(DOB)引发的rRNA基因的突变情况,本研究在DOB基因组De novo测序的基础上应用生物信息学方法分析了3株离子束重组DOB菌株DOB073、DOB113、DOB981与原始菌株DOB150的rRNA基因序列,并对其rRNA基因的突变进行了研究。通过从DOB基因组De novo测序数据中获取这些DOB菌株的5srRNA、16S rRNA、23srRNA的基因序列,研究其基因突变的基本规律,分析基因突变的具体位点,统计其拷贝数、GC含量、计算进化距离和分子钟,进而构建进化树,并分析基因的进化程度,寻找并分析比对结果中的保守区域。rRNA基因拷贝数分析结果表明:重组突变菌DOB073的5srRNA和16s rRNA基因拷贝数分别比原始菌株DOB150增加了14和5个;重组突变菌DOB113的16s rRNA基因拷贝数比原始菌株DOB150增加了5个;重组突变菌DOB981的5srRNA基因拷贝数比原始菌株DOB150减少了1个,而其16s rRNA基因拷贝数增加了5个;3株重组突变菌株的23s rRNA基因拷贝数与对照菌株DOB150相同。rRNA基因碱基突变位点的研究结果显示,3株重组突变菌的5srRNA基因均在第4、107、112、114处发生了碱基突变,DOB073和DOB113还在第104处发生了碱基突变;3株重组突变菌的16srRNA基因的5-6个碱基突变分别分布于C1区、V1区、V2区、V4区、V6区、V7和V9区;3株重组突变菌的23srRNA基因的保守性较强,只有2-3个碱基位点发生了突变。基因进化分析结果显示,重组突变菌株DOB073和DOB113的5srRNA基因和23srRNA基因的进化速度均快于重组突变菌株DOB981。而DOB981的16srRNA基因的进化速度快于DOB073和DOB113。rRNA基因序列的保守二级结构的研究结果表明:三株重组突变菌株的16srRNA基因和原始菌株的16srRNA基因均具有9个保守二级结构,其碱基位置分别为:80-120,120-240,240-360,360-480,400-520,640-760,760-880,800-920和1420-1540。而三株重组突变菌株的23srRNA基因和原始菌株的23srRNA基因都均具有13个保守二级结构,其处碱基位置分别为:40-160,80-200,120-240,720-840,1200-1320,1480-1600,1520-1640,2120-2240,2200-2320,2240-2360,2280-2400,2360-2480,2520-2640。通过对3株重组突变DOB菌株的3种rRNA基因的研究,从分子水平上揭示了离子注入介导DOB菌株rRNA基因突变与进化的分子机制,也为离子注入介导获得的DOB的多样性提供了理论和实践依据,特别是是为低能离子注入介导的原核微生物的进化提供了直接的分子证据。
[Abstract]:Low energy ion implantation is a physical mutagenic breeding technique developed in recent years. Thus triggering biological effects. In order to understand the mutation of rRNA gene induced by oligonutrient desert bacteria (DOB) mediated by low energy N injection, On the basis of DOB genomic De novo sequencing, the rRNA gene sequences of three ion-beam recombinant DOB strains DOB073,DOB113,DOB981 and the original DOB150 were analyzed by bioinformatics, and their rRNA gene mutations were studied. The gene sequence of 5srRNA-16s rRNA,23srRNA of these DOB strains was obtained from DOB genomic De novo sequencing data, the basic rules of gene mutation were studied, the specific sites of gene mutation were analyzed, and the copy number and GC content were counted. Calculate the evolutionary distance and molecular clock, and then build the evolutionary tree, and analyze the degree of evolution of the gene. The copy number analysis of rRNA gene showed that the copy number of 5srRNA gene and 16s rRNA gene of recombinant mutant DOB073 were 14 and 5 more than that of the original strain DOB150, respectively. The copy number of 16s rRNA gene of recombinant mutant DOB113 was 5 more than that of original strain DOB150, the copy number of 5srRNA gene of recombinant mutant DOB981 was 1 less than that of original strain DOB150, and the copy number of 16s rRNA gene increased by 5. The copy number of 23s rRNA gene of three recombinant mutant strains was the same as that of control strain DOB150. The results of the study on the base mutation site of rRNA gene showed that the 5srRNA gene of the three recombinant mutants had a base mutation at 4107112114. DOB073 and DOB113 also had base mutation at 104th. The 5-6 base mutations of 16srRNA gene of the three recombinant mutant strains were located in C1 region, V1 region, V2 region, V4 region, V6 region, V7 and V9 region, respectively. The 23srRNA gene of the three recombinant mutants was highly conserved, and only 2-3 base loci were mutated. The results of gene evolution analysis showed that the 5srRNA gene and 23srRNA gene of the recombinant mutant DOB073 and DOB113 were both faster than that of the recombinant mutant DOB981.. The results showed that the 16srRNA gene of the three recombinant mutant strains and the 16srRNA gene of the original strain had 9 conserved secondary structures, while the evolution rate of the 16srRNA gene of DOB981 was faster than that of the conserved secondary structure of the DOB073 and DOB113.rRNA gene sequences. Their base positions are 80-120120-240240-360360-480400-520640-760760-880800-920 and 1420-1540 respectively. The 23srRNA gene of the three recombinant mutants and the 23srRNA gene of the original strain all had 13 conserved secondary structures, and their base positions were as follows: 40-160 (80-200120-240720-84040) 1200-1320 (1480-1600) 1520-164040 (2120-2240) 2200-2320 (2240-23602280-2400) 2360-2480 (2520-2640). Through the study of three rRNA genes of three recombinant mutant DOB strains, the molecular mechanism of rRNA gene mutation and evolution mediated by ion implantation was revealed at the molecular level. It also provides a theoretical and practical basis for the diversity of DOB mediated by ion implantation, especially provides direct molecular evidence for the evolution of prokaryotic microorganisms mediated by low energy ion implantation.
【学位授予单位】：新疆大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q78
本文编号：2388415
[Abstract]:Low energy ion implantation is a physical mutagenic breeding technique developed in recent years. Thus triggering biological effects. In order to understand the mutation of rRNA gene induced by oligonutrient desert bacteria (DOB) mediated by low energy N injection, On the basis of DOB genomic De novo sequencing, the rRNA gene sequences of three ion-beam recombinant DOB strains DOB073,DOB113,DOB981 and the original DOB150 were analyzed by bioinformatics, and their rRNA gene mutations were studied. The gene sequence of 5srRNA-16s rRNA,23srRNA of these DOB strains was obtained from DOB genomic De novo sequencing data, the basic rules of gene mutation were studied, the specific sites of gene mutation were analyzed, and the copy number and GC content were counted. Calculate the evolutionary distance and molecular clock, and then build the evolutionary tree, and analyze the degree of evolution of the gene. The copy number analysis of rRNA gene showed that the copy number of 5srRNA gene and 16s rRNA gene of recombinant mutant DOB073 were 14 and 5 more than that of the original strain DOB150, respectively. The copy number of 16s rRNA gene of recombinant mutant DOB113 was 5 more than that of original strain DOB150, the copy number of 5srRNA gene of recombinant mutant DOB981 was 1 less than that of original strain DOB150, and the copy number of 16s rRNA gene increased by 5. The copy number of 23s rRNA gene of three recombinant mutant strains was the same as that of control strain DOB150. The results of the study on the base mutation site of rRNA gene showed that the 5srRNA gene of the three recombinant mutants had a base mutation at 4107112114. DOB073 and DOB113 also had base mutation at 104th. The 5-6 base mutations of 16srRNA gene of the three recombinant mutant strains were located in C1 region, V1 region, V2 region, V4 region, V6 region, V7 and V9 region, respectively. The 23srRNA gene of the three recombinant mutants was highly conserved, and only 2-3 base loci were mutated. The results of gene evolution analysis showed that the 5srRNA gene and 23srRNA gene of the recombinant mutant DOB073 and DOB113 were both faster than that of the recombinant mutant DOB981.. The results showed that the 16srRNA gene of the three recombinant mutant strains and the 16srRNA gene of the original strain had 9 conserved secondary structures, while the evolution rate of the 16srRNA gene of DOB981 was faster than that of the conserved secondary structure of the DOB073 and DOB113.rRNA gene sequences. Their base positions are 80-120120-240240-360360-480400-520640-760760-880800-920 and 1420-1540 respectively. The 23srRNA gene of the three recombinant mutants and the 23srRNA gene of the original strain all had 13 conserved secondary structures, and their base positions were as follows: 40-160 (80-200120-240720-84040) 1200-1320 (1480-1600) 1520-164040 (2120-2240) 2200-2320 (2240-23602280-2400) 2360-2480 (2520-2640). Through the study of three rRNA genes of three recombinant mutant DOB strains, the molecular mechanism of rRNA gene mutation and evolution mediated by ion implantation was revealed at the molecular level. It also provides a theoretical and practical basis for the diversity of DOB mediated by ion implantation, especially provides direct molecular evidence for the evolution of prokaryotic microorganisms mediated by low energy ion implantation.
【学位授予单位】：新疆大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q78
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