利用16S-23S rRNA ITS AFLP指纹图谱技术监控四川传统米酒发酵过程中原核微生物的演替,并结合米酒发酵过程中理化因子的动态变化对其演替过程进行了分析。研究结果表明,米酒发酵过程中,随着酒曲的接入,米酒醅中原核微生物伴随米酒理化...利用16S-23S rRNA ITS AFLP指纹图谱技术监控四川传统米酒发酵过程中原核微生物的演替,并结合米酒发酵过程中理化因子的动态变化对其演替过程进行了分析。研究结果表明,米酒发酵过程中,随着酒曲的接入,米酒醅中原核微生物伴随米酒理化因子的动态变化而发生群落演替。在适应期和发酵期,米酒醅中的原核微生物群落分别聚成群I和群II,二者相关系数为0.49。群II中,在相关系数0.638水平上又分为IIA和IIB两个分支。分支IIA又进一步分为IIA1和IIA2两簇,相关系数为0.73。簇IIA2中主要发生的是发酵旺盛期微生物的演替;簇IIA1中I,IA1-1和IIA1-2亚簇分别表示发酵前期、发酵后期米酒醅中的原核微生物群落演替,二者聚集于相关系数0.80。其中I,IA1-2中,发酵52 h和55 h的米酒醅中原核微生物群落几乎相似,二者群落相关系数为1.0。展开更多
目的利用16 s-23 s rRNA间隔区(ITS)的多态性,对中国布鲁氏菌种间或种内生物型进行鉴别,评价ITS作为基因标识物的意义,寻找适合布鲁氏菌分型研究的基因标识物。方法应用聚合酶链反应-单链构象多态性(PCR-SSCP)分析技术,对中国120株布鲁...目的利用16 s-23 s rRNA间隔区(ITS)的多态性,对中国布鲁氏菌种间或种内生物型进行鉴别,评价ITS作为基因标识物的意义,寻找适合布鲁氏菌分型研究的基因标识物。方法应用聚合酶链反应-单链构象多态性(PCR-SSCP)分析技术,对中国120株布鲁氏菌的ITS进行分析和筛选,测序结果与Genbank中的布鲁氏菌ITS序列进行比较分析。结果对16 s-23 s rRNA间隔区的SSCP结果分析,得到4种不同的带型(ⅠI、I、Ⅲ、Ⅳ),测序序列有3个位点的差异,达到99.87%的一致性。结论中国布鲁氏菌的ITS序列高度保守,具有一定探讨其作为布鲁氏菌属种内分型的基因标识的价值。展开更多
The phylogeny of γ-proteobacteria was inferred from nucleotide sequence comparisons of a short 232 nucleotide sequence marker. A total of 64 γ-proteobacterial strains from 13 Orders, 22 families, 40 genera and 59 sp...The phylogeny of γ-proteobacteria was inferred from nucleotide sequence comparisons of a short 232 nucleotide sequence marker. A total of 64 γ-proteobacterial strains from 13 Orders, 22 families, 40 genera and 59 species were analyzed. The short 232 nucleotide sequence marker used here was a combination of a 157 nucleotide sequence at the 3’ end of the 16S rRNA gene and a 75 nucleotide sequence at the 5’ end of the 16S-23S Internal Transcribed Spacer (ITS) sequence. Comparative analyses of the 3’ end of the 16S rRNA gene nucleotide sequence showed that the last 157 bp were conserved among strains from same species and less conserved in more distantly related species. This 157 bp sequence was selected as the first part in the construction of our nucleotide sequence marker. A bootstrapped neighbor-joining tree based on the alignment of this 157 bp was constructed. This 157 bp could distinguish γ-proteobacterial species from different genera from same family. Closely related species could not be distinguished. Next, an alignment of the 16S-23S ITS nucleotide sequences of alleles from same bacterial strain was performed. The first 75 bp at the 5’ end of the 16S-23S ITS was highly conserved at the intra-strain level. It was selected as the second part in the construction of our nucleotide sequence marker. Finally, a bootstrapped neighbor-joining tree based on the alignment of this 232 bp sequence was constructed. Based on the topology of the neighbour-joining tree, four major Groups, Group I to IV, were revealed with several sub-groups and clusters. Our results, based on the 232 bp sequence were, in general, in agreement with the phylogeny of γ-proteobacteria based on the 16S rRNA gene. The use of this 232 bp sequence as a phylogenetic marker presents several advantages over the use of the entire 16S rRNA gene or the generation of extensive phenotypic and genotypic data in phylogenetic analyses. First, this marker is not allele-dependant. Second, this 232 bp marker contains 157 bp from the 3’ end of the 16S rRNA gene and 75 bp from the 5’ end of the 16S-23S ITS. The 157 bp allows discrimination among distantly related species. Owing to its higher rate of nucleotide substitutions, the 75 bp adds discriminating power among closely related species from same genus and closely related genera from same family. Because of its higher percentage of nucleotide sequence divergence than the 16S rRNA gene, the 232 bp marker can better discriminate among closely related γ-proteobacterial species. Third, the method is simple, rapid, suited to large screening programs and easily accessible to most laboratories. Fourth, this marker can also reveal γ-proteobacterial species which may appear misassigned and for which additional characterization appear warranted.展开更多
文摘目的利用16 s-23 s rRNA间隔区(ITS)的多态性,对中国布鲁氏菌种间或种内生物型进行鉴别,评价ITS作为基因标识物的意义,寻找适合布鲁氏菌分型研究的基因标识物。方法应用聚合酶链反应-单链构象多态性(PCR-SSCP)分析技术,对中国120株布鲁氏菌的ITS进行分析和筛选,测序结果与Genbank中的布鲁氏菌ITS序列进行比较分析。结果对16 s-23 s rRNA间隔区的SSCP结果分析,得到4种不同的带型(ⅠI、I、Ⅲ、Ⅳ),测序序列有3个位点的差异,达到99.87%的一致性。结论中国布鲁氏菌的ITS序列高度保守,具有一定探讨其作为布鲁氏菌属种内分型的基因标识的价值。
文摘The phylogeny of γ-proteobacteria was inferred from nucleotide sequence comparisons of a short 232 nucleotide sequence marker. A total of 64 γ-proteobacterial strains from 13 Orders, 22 families, 40 genera and 59 species were analyzed. The short 232 nucleotide sequence marker used here was a combination of a 157 nucleotide sequence at the 3’ end of the 16S rRNA gene and a 75 nucleotide sequence at the 5’ end of the 16S-23S Internal Transcribed Spacer (ITS) sequence. Comparative analyses of the 3’ end of the 16S rRNA gene nucleotide sequence showed that the last 157 bp were conserved among strains from same species and less conserved in more distantly related species. This 157 bp sequence was selected as the first part in the construction of our nucleotide sequence marker. A bootstrapped neighbor-joining tree based on the alignment of this 157 bp was constructed. This 157 bp could distinguish γ-proteobacterial species from different genera from same family. Closely related species could not be distinguished. Next, an alignment of the 16S-23S ITS nucleotide sequences of alleles from same bacterial strain was performed. The first 75 bp at the 5’ end of the 16S-23S ITS was highly conserved at the intra-strain level. It was selected as the second part in the construction of our nucleotide sequence marker. Finally, a bootstrapped neighbor-joining tree based on the alignment of this 232 bp sequence was constructed. Based on the topology of the neighbour-joining tree, four major Groups, Group I to IV, were revealed with several sub-groups and clusters. Our results, based on the 232 bp sequence were, in general, in agreement with the phylogeny of γ-proteobacteria based on the 16S rRNA gene. The use of this 232 bp sequence as a phylogenetic marker presents several advantages over the use of the entire 16S rRNA gene or the generation of extensive phenotypic and genotypic data in phylogenetic analyses. First, this marker is not allele-dependant. Second, this 232 bp marker contains 157 bp from the 3’ end of the 16S rRNA gene and 75 bp from the 5’ end of the 16S-23S ITS. The 157 bp allows discrimination among distantly related species. Owing to its higher rate of nucleotide substitutions, the 75 bp adds discriminating power among closely related species from same genus and closely related genera from same family. Because of its higher percentage of nucleotide sequence divergence than the 16S rRNA gene, the 232 bp marker can better discriminate among closely related γ-proteobacterial species. Third, the method is simple, rapid, suited to large screening programs and easily accessible to most laboratories. Fourth, this marker can also reveal γ-proteobacterial species which may appear misassigned and for which additional characterization appear warranted.
