为了解烟草根部内生细菌多样性,采用改进的CTAB法提取烟草根部总DNA,利用细菌16 S rDNA基因通用引物对烟草根总DNA进行16 S rDNA扩增,构建烟草根部内生细菌16 S rDNA克隆文库;挑取具有不同酶切谱型的克隆进行测序、比对并构建16 S rDNA...为了解烟草根部内生细菌多样性,采用改进的CTAB法提取烟草根部总DNA,利用细菌16 S rDNA基因通用引物对烟草根总DNA进行16 S rDNA扩增,构建烟草根部内生细菌16 S rDNA克隆文库;挑取具有不同酶切谱型的克隆进行测序、比对并构建16 S rDNA基因系统发育树。构建的烟草根部内生细菌16 S rDNA克隆文库中,155个克隆分属于36个不同的分类单元,Blast结果表明,大部分克隆与已知细菌的16 S rDNA序列相似性较高,分别属于变形菌门(Proteobacteria)的Alpha、Gamma、Beta亚群,放线菌门(Actinobacteria),拟杆菌门(Bacteroidetes),厚壁菌门(Fir-micutes)中的肠杆菌属(Enterobacter)、不动杆菌属(Acinetobacter)、寡养食单胞菌属(Stenotrophomonas)、假单胞菌属(Pseudomonas)、嗜甲基菌属(Methylobacillus)、食酸菌属(Acidovorax)、芽孢杆菌属(Bacillus)等16个属,其中13.5%的克隆序列与非可培养细菌(Uncultured bacteria)的相似性在93%～99%之间,假单胞菌属细菌是烟草根部内生细菌的优势种群。这些研究结果说明烟草根部存在较为丰富的内生细菌系统发育多样性,并且潜藏着未知的内生细菌资源。展开更多
Natural products derived from bacterial sources have long been pivotal in the discovery of drug leads.However,the cultivation of only about 1%of bacteria in laboratory settings has left a significant portion of biosyn...Natural products derived from bacterial sources have long been pivotal in the discovery of drug leads.However,the cultivation of only about 1%of bacteria in laboratory settings has left a significant portion of biosynthetic diversity hidden within the genomes of uncultured bacteria.Advances in sequencing technologies now enable the exploration of genetic material from these metagenomes through culture-independent methods.This approach involves extracting genetic sequences from environmental DNA and applying a hybrid methodology that combines functional screening,sequence tag-based homology screening,and bioinformatic-assisted chemical synthesis.Through this process,numerous valuable natural products have been identified and synthesized from previously uncharted metagenomic territories.This paper provides an overview of the recent advancements in the utilization of culture-independent techniques for the discovery of novel biosynthetic gene clusters and bioactive small molecules within metagenomic libraries.展开更多
Increasing attention has been paid to phosphate-accumulating organisms （PAOs） for their important role in biological phosphorus removal. In this study, microbial communities of PAOs cultivated under different carbon...Increasing attention has been paid to phosphate-accumulating organisms （PAOs） for their important role in biological phosphorus removal. In this study, microbial communities of PAOs cultivated under different carbon sources （sewage, glucose, and sodium acetate） were investigated and compared through culture-dependent and culture-independent methods, respectively. The results obtained using denaturing gradient gel electrophoresis （DGGE） of polymerase chain reaction-amplified 16S rDNA fragments revealed that the diversity of bacteria in a sewage-fed reactor （1#） was much higher than in a glucose-fed one （2#） and a sodium acetate-fed one （3#）; there were common PAOs in three reactors fed by different carbon sources. Five strains were separated from three systems by using a phosphaterich medium; they were from common bacteria isolated and three isolates could not be found in DGGE profile at all. Two isolates had good phosphorus removal ability. When the microbial diversity was studied, the molecular biological method was better than the culture-dependent one. When phosphorus removal characteristics were investigated, culture-dependent approach was more effective. Thus a combination of two methods is necessary to have a comprehensive view of PAOs.展开更多
More than 99% of identified prokaryotes, including many from the marine environment, cannot be cultured in the laboratory. This lack of capability restricts our knowledge of microbial genetics and community ecology. M...More than 99% of identified prokaryotes, including many from the marine environment, cannot be cultured in the laboratory. This lack of capability restricts our knowledge of microbial genetics and community ecology. Metagenomics, the culture-independent cloning of environmental DNAs that are isolated directly from an environmental sample, has already provided a wealth of information about the uncultured microbial world. It has also facilitated the discovery of novel bio- catalysts by allowing researchers to probe directly into a huge diversity of enzymes within natural microbial communities. Recent advances in these studies have led to a great interest in recruiting microbial enzymes for the development of environmentally-friendly industry. Although the metage- nomics approach has many limitations, it is expected to provide not only scientific insights but also economic benefits, especially in industry. This review highlights the importance of metagenomics in mining microbial lipases, as an example, by using high-throughput techniques. In addition, we dis- cuss challenges in the metagenomics as an important part of bioinformatics analysis in big data.展开更多
【目的】免培养和纯培养相结合分析南海深海沉积物放线菌多样性。【方法】免培养方法通过提取沉积物宏基因组DNA,利用放线菌门特异性引物扩增放线菌16S r RNA基因序列,构建放线菌16S r RNA基因克隆文库,文库经RFLP(Restriction fragment...【目的】免培养和纯培养相结合分析南海深海沉积物放线菌多样性。【方法】免培养方法通过提取沉积物宏基因组DNA,利用放线菌门特异性引物扩增放线菌16S r RNA基因序列,构建放线菌16S r RNA基因克隆文库,文库经RFLP(Restriction fragment length polymorphism)分析后挑选代表序列测序并进行多样性指数分析和系统发育分析。可培养方法利用8种培养基进行菌株分离,对排重后的菌株进行16S r RNA基因序列多样性分析。【结果】构建的两个深海位点的16S r RNA基因克隆文库在放线菌门的放线菌纲(Actinobacteria)、酸微菌纲(Acidimicrobiia)、腈基降解菌纲(Nitriliruptoria)和嗜热油菌纲(Thermoleophilia)4个纲中均有分布;两个位点中的种群结构有差异,N40-4位点的优势种群是放线菌纲的链霉菌目(Streptomycetales);N63-4位点的优势种群是腈基降解菌纲的腈基降解菌目(Nitriliruptorales)。8种培养基共分离出41株放线菌,根据形态特征排重后得到的19株菌分布于10个不同的属,12个不同的种,其中稀有放线菌属比例较高,菌株OAct400为潜在的微杆菌属(Microbacterium)新种。【结论】南海深海沉积物蕴含着丰富的放线菌物种资源及大量未知种群,具有进一步研究的价值。展开更多
文摘为了解烟草根部内生细菌多样性,采用改进的CTAB法提取烟草根部总DNA,利用细菌16 S rDNA基因通用引物对烟草根总DNA进行16 S rDNA扩增,构建烟草根部内生细菌16 S rDNA克隆文库;挑取具有不同酶切谱型的克隆进行测序、比对并构建16 S rDNA基因系统发育树。构建的烟草根部内生细菌16 S rDNA克隆文库中,155个克隆分属于36个不同的分类单元,Blast结果表明,大部分克隆与已知细菌的16 S rDNA序列相似性较高,分别属于变形菌门(Proteobacteria)的Alpha、Gamma、Beta亚群,放线菌门(Actinobacteria),拟杆菌门(Bacteroidetes),厚壁菌门(Fir-micutes)中的肠杆菌属(Enterobacter)、不动杆菌属(Acinetobacter)、寡养食单胞菌属(Stenotrophomonas)、假单胞菌属(Pseudomonas)、嗜甲基菌属(Methylobacillus)、食酸菌属(Acidovorax)、芽孢杆菌属(Bacillus)等16个属,其中13.5%的克隆序列与非可培养细菌(Uncultured bacteria)的相似性在93%～99%之间,假单胞菌属细菌是烟草根部内生细菌的优势种群。这些研究结果说明烟草根部存在较为丰富的内生细菌系统发育多样性,并且潜藏着未知的内生细菌资源。
基金supported by the Start-Up Grant from China Pharmaceutical University(No.3154070046).
文摘Natural products derived from bacterial sources have long been pivotal in the discovery of drug leads.However,the cultivation of only about 1%of bacteria in laboratory settings has left a significant portion of biosynthetic diversity hidden within the genomes of uncultured bacteria.Advances in sequencing technologies now enable the exploration of genetic material from these metagenomes through culture-independent methods.This approach involves extracting genetic sequences from environmental DNA and applying a hybrid methodology that combines functional screening,sequence tag-based homology screening,and bioinformatic-assisted chemical synthesis.Through this process,numerous valuable natural products have been identified and synthesized from previously uncharted metagenomic territories.This paper provides an overview of the recent advancements in the utilization of culture-independent techniques for the discovery of novel biosynthetic gene clusters and bioactive small molecules within metagenomic libraries.
基金Project supported by the Young Teacher Foundation of Donghua University(No.113-10-0044065)Key(keygrant)Project of Chinese Ministry of Education(No.107046)Program for New Century Excellent Talents in University(NCET)
文摘Increasing attention has been paid to phosphate-accumulating organisms （PAOs） for their important role in biological phosphorus removal. In this study, microbial communities of PAOs cultivated under different carbon sources （sewage, glucose, and sodium acetate） were investigated and compared through culture-dependent and culture-independent methods, respectively. The results obtained using denaturing gradient gel electrophoresis （DGGE） of polymerase chain reaction-amplified 16S rDNA fragments revealed that the diversity of bacteria in a sewage-fed reactor （1#） was much higher than in a glucose-fed one （2#） and a sodium acetate-fed one （3#）; there were common PAOs in three reactors fed by different carbon sources. Five strains were separated from three systems by using a phosphaterich medium; they were from common bacteria isolated and three isolates could not be found in DGGE profile at all. Two isolates had good phosphorus removal ability. When the microbial diversity was studied, the molecular biological method was better than the culture-dependent one. When phosphorus removal characteristics were investigated, culture-dependent approach was more effective. Thus a combination of two methods is necessary to have a comprehensive view of PAOs.
基金supported by King Abdullah University of Science and Technology (KAUST),Saudi Arabia
文摘More than 99% of identified prokaryotes, including many from the marine environment, cannot be cultured in the laboratory. This lack of capability restricts our knowledge of microbial genetics and community ecology. Metagenomics, the culture-independent cloning of environmental DNAs that are isolated directly from an environmental sample, has already provided a wealth of information about the uncultured microbial world. It has also facilitated the discovery of novel bio- catalysts by allowing researchers to probe directly into a huge diversity of enzymes within natural microbial communities. Recent advances in these studies have led to a great interest in recruiting microbial enzymes for the development of environmentally-friendly industry. Although the metage- nomics approach has many limitations, it is expected to provide not only scientific insights but also economic benefits, especially in industry. This review highlights the importance of metagenomics in mining microbial lipases, as an example, by using high-throughput techniques. In addition, we dis- cuss challenges in the metagenomics as an important part of bioinformatics analysis in big data.
文摘【目的】免培养和纯培养相结合分析南海深海沉积物放线菌多样性。【方法】免培养方法通过提取沉积物宏基因组DNA,利用放线菌门特异性引物扩增放线菌16S r RNA基因序列,构建放线菌16S r RNA基因克隆文库,文库经RFLP(Restriction fragment length polymorphism)分析后挑选代表序列测序并进行多样性指数分析和系统发育分析。可培养方法利用8种培养基进行菌株分离,对排重后的菌株进行16S r RNA基因序列多样性分析。【结果】构建的两个深海位点的16S r RNA基因克隆文库在放线菌门的放线菌纲(Actinobacteria)、酸微菌纲(Acidimicrobiia)、腈基降解菌纲(Nitriliruptoria)和嗜热油菌纲(Thermoleophilia)4个纲中均有分布;两个位点中的种群结构有差异,N40-4位点的优势种群是放线菌纲的链霉菌目(Streptomycetales);N63-4位点的优势种群是腈基降解菌纲的腈基降解菌目(Nitriliruptorales)。8种培养基共分离出41株放线菌,根据形态特征排重后得到的19株菌分布于10个不同的属,12个不同的种,其中稀有放线菌属比例较高,菌株OAct400为潜在的微杆菌属(Microbacterium)新种。【结论】南海深海沉积物蕴含着丰富的放线菌物种资源及大量未知种群,具有进一步研究的价值。
