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7, Effect of storage conditions on the assessment of bacterial community structure in soil and human-associated samples. Â
13. Validation of heavy-water stable isotope probing for the characterization of rapidly responding soil bacteria. Â
14. Identification of Nitrogen-Incorporating Bacteria in Petroleum-Contaminated Arctic Soils Using 15N DNA-SIP and Pyrosequencing. Â
17, Disclosing arbuscular mycorrhizal fungal biodiversity in soil through a land-use gradient using a pyrosequencing approach. Â
20, Tag-encoded pyrosequencing analysis of bacterial diversity in a single soil type as affected by management and land use.
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454 High-throughput Sequencing - A New Approach to Soil Microorganisms
In terrestrial ecosystems, there are large numbers of microbial populations in the soil, including prokaryotic microorganisms such as bacteria, cyanobacteria, actinomycetes and ultramicroscopic microorganisms, as well as eukaryotes such as fungi, algae (except cyanobacteria), and lichens. Wait. They have a clear division of labor with plants and animals, mainly playing the role of “decomposersâ€, and are involved in almost all biological and biochemical reactions in the soil, and are responsible for the “regulators†of the Earth's C, N, P, S and other material cycles [1] ] , the “transformer†of soil nutrient plant availability and the “purifier†of polluting the environment, etc. [2] . Soil microbes are sensitive indicators of climate and soil environmental conditions. Soil microbial community structure and diversity and their changes reflect soil quality to some extent. Soil microbial diversity generally includes microbial taxa diversity, genetic (gene) diversity, ecological diversity and functional diversity [3] . Due to the complexity of soil microbes, the variability of the soil itself, and the imperfect research methods, the research on soil microbial diversity has been far behind in comparison with animals and plants. With the rapid development of modern biological molecular biology techniques such as polymerase chain reaction (PCR) and nucleic acid sequencing, people have more understanding of soil microbial diversity; the development of high-throughput sequencing technology provides research for soil metagenomics. A large amount of data provides objective and comprehensive information for directly exploring the microbial community structure in the soil.
1. Methods for studying soil microorganisms
1. Microbial plate culture method
Most of the microbial community diversity and structural analysis in traditional soil ecosystems are isolated and cultured, and then analyzed by general biochemical traits or specific phenotypes, limited to the separation of microorganisms from solid media.
This method is limited to a very small amount (0.1%-1%) of microbial populations that can be cultured, and it is impossible to conduct an in-depth study on the taxonomic status and phylogenetic relationships of most microorganisms.
2, molecular biology technology combined with sequencing methods
In the past 20 years, molecular biology techniques, especially 16SrDNA technology, have been widely used in the identification of unknown bacteria. Since the 1980s, research methods for modern microbial molecular ecology based on molecular phylogenetic analysis have been gradually established, such as PCR-RFLP, PCR-RAPD, PCR-SSCP, fluorescence in situ hybridization (FISH), and genes. Microarry, Phospholipid fatty acid (PLFA), Stable Isotope Probing (SIP), PCR-DGGE/TGGE (Denaturing gradient gel electrophoresis, DGGE/Temperature gradient gel electrophoresis, TGGE) , enabling researchers to study soil microbial diversity at the molecular level.
Among them, the PCR-DGGE/TGGE method is widely used and widely accepted. The limitation of DGGE/TGGE is that the length of the detected DNA fragments ranges from 200 bp to 900 bp. Fragments outside this range are difficult to detect [4] , and the G/C base pair required for PCR amplification is at least 40%. Only the dominant flora in the environment can be detected. Only the group that accounts for about 1% or more of the total number of bacteria in the community can be detected by DGGE [5] . The temperature range allowed by TGGE instruments ranges from 15 °C to 80 °C. The Tm value of larger fragment DNA is more difficult to detect because it is close to 80 °C. If the electrophoresis conditions are not suitable, the DNA fragments with sequence differences cannot be completely guaranteed. A phenomenon in which DNAs with different sequences migrate in the same position.
3. High-throughput sequencing method
In recent years, 16S rRNA/DNA-based molecular biology techniques have become a widely accepted method [6,7] . Studies have shown that 400-600 base sequences are sufficient for preliminary estimation of microbial diversity and population classification in the environment [8] , so the 454 high-throughput sequencing method is long and accurate due to its read length (400~500bp). Highly characterized features are used extensively for the study of microbial diversity.
With the 16S rDNA sequence of the microorganism, both full length and part, it can be submitted to GenBank for similarity analysis using the BLAST program and known sequences. Gen Bank will list the known sequence lists, the degree of similarity, and the microbial species corresponding to these sequences, based on the similarity to the measured sequences, but more precise microbial classification depends on phylogenetic analysis.
The superiority of high-throughput sequencing is as follows: the sequencing sequence is long, covering high-variation regions such as 16S / 18S rDNA and ITS; high sequencing flux can detect trace microorganisms in environmental samples; the experiment is simple and the results are stable. Reproducible; no need for complex library construction, microbial DNA amplification products can be directly sequenced, the experimental cycle is short; sequencing data is convenient for bioinformatics analysis.
This method has been recognized by top journals (Nature, etc.) and has become an important means of soil microbial diversity detection. Shanghai Meiji Biotech has had several successful cases to conduct high-throughput sequencing and bioinformatics analysis of soil samples provided by customers. Because of its large amount of data and simple operation process, high-throughput sequencing avoids the sample loss caused by the tedious experimental operation as much as possible, and can reflect the actual situation of the soil sample relatively objectively.
Second, the application of high-throughput sequencing in soil microbial research
1.1 Studying the species diversity of soil microbes
Microbial species diversity mainly consists of the number and proportion of microbial groups, namely bacteria, fungi and actinomycetes, to describe microbial diversity, or to divide them into different functional groups according to their role in ecosystems. (function group), study the soil microbial diversity through the classification and quantity of species in a functional group, such as the diversity of methanogenic bacteria, nitrogen-fixing bacteria, rhizobium in soil. Here are a few representative articles.
Roesch et al [9] used the 454 sequencing method to detect and statistically evaluate a large cross-section of four types of soil in the Western Hemisphere. The results showed that among the four soils, the most abundant microbial group was Bacteroides, β-Proteobacteria and α-Proteobacteria. Compared with agricultural soils, the soil microbial diversity is more abundant. However, the results show that the archaea diversity in forest soil is less, only 0.009% of all sequences in this locus, while the proportion of agricultural soil is 4% -12%.
The diversity of bacteria in the soil is very large, and the bacterial population in different soils is also diversified. Triplett et al [10] sequenced the V2-V3 region of 16S rRNA based on pyrosequencing to estimate the overall and vertical characteristics of the flora in nine grassland soils. Cluster analysis was performed on all 752,838 data sequences to explore the specificity of the flora in terms of abundance, diversity and composition. The authors found that in different soil layers, different distributions of populations or subpopulations of bacterial systems are related to soil properties, including organic carbon content, total nitrogen levels, or microbial biomass.
1.2 Studying soil microbial functional diversity
Soil microbial functional diversity refers to microbial activity, substrate metabolism, and functions related to the transformation of nutrients such as N, P, and S in the soil. Some transformation processes in the soil, such as organic carbon, nitrification, and The activity of enzymes in the soil, etc., to understand soil microbial function.
Ammoxidation is the first step in nitrification and an important process in the global nitrogen cycle for the formation of nitrifying salts by microbial activity. Leininger [11] examined the gene abundance of a subunit encoding ammonia monooxygenase (amoA) in 12 primary and agricultural land in three climate regions. Reverse transcription quantitative PCR and non-cloning pyrosequencing techniques were used to sequence complementary DNA, confirming that the ammoxidation activity of archaea is much higher than that of bacteria, confirming that Crenarchaeota may be the most abundant ammonia-oxidizing microorganism in soil ecosystem.
Urich et al. [12] used RNA-based environmental transcriptomics to simultaneously obtain soil microbial population structure and function information. The results show that this method can simultaneously study the structure and function of microbial populations to avoid deviations caused by other methods. Community genomics analysis can obtain information on microbial function by studying the relationship between microbial genome sequences and certain expression characteristics. But there are other ways to map specific functions to microbial communities with this specific function. Quantification and comparative analysis of genes encoding rRNA-expressing genes and major enzymes related to environmental factors will enable understanding of the relationship between microbial structure and specific functions, such as nitrification, denitrification, and contaminant degradation.
Denitrification is one of the important processes involved in the nitrogen cycle, such as nitrogen loss and greenhouse gas emissions. Through macrogenomic sequencing, combined with molecular detection and pyrosequencing, Ryan et al. isolated and manipulated the enzymes encoding the denitrification process. By screening the clones obtained from 77,000 soil metagenomic libraries, nine enzyme clusters involved in denitrification were finally isolated and identified [13] .
1.3 Impact of sudden changes in the research environment on soil microbial flora
Sudden changes in the environment can cause changes in the structure and function of the microbial community. Zachary et al. [14] identified bacteria associated with soil humidification with heavy water stable isotope detection technology (H218O-SIP). By liquid chromatography/mass spectrometry (LC-MS), the authors determined that the oxygen atoms in H218O bind to all of the DNA structural components. Although this binding is not uniform, it is still possible to distinguish between the labeled 18O and the unlabeled DNA. The authors found that the oxygen atoms in the DNA and extracellular H2O did not exchange in vitro, indicating that the 18O incorporated into the DNA is relatively stable and the proportion of 18O incorporated into the bacterial DNA is higher (48-72 h). After soil humidification, high-throughput sequencing of 16S rRNA in soil showed that the relative proportions of Alphaproteobacteria , Betaproteobacteria and Gammaproteobacteria increased, while the ratio of Chloroflexi and Deltapopoteobacteria decreased. By controlling the dynamic changes of soil moisture, the author studies the structural changes of microbial flora and the division of ecological groups.
Note: Summary of articles published using 454 pyrosequencing
1. Archaea predominate among ammonia-oxidizing prokaryotes in soils
—— (2006) Nature 442: 806-809 - if 36.101
2, 454 Pyrosequencing analyses of forest soils reveal an unexpectedly high fungal diversity. Â
——(2009) New Phytologist 184(2): 449-56.—— if 6.516
3, Vertical distribution of fungal communities in tallgrass prairie soil.
——(2010) Mycologia 102(5): 1027-41.—— if 6.2
4, A survey of soil acidobacterial diversity using pyrosequencing and clone library analyses. Â
——(2009) ISME Journal 3(4): 442-53—— if 6.153
5, Pyrosequencing enumerates and contrasts soil microbial diversity
——(2008) The ISME Journal 1: 283–290 – if 6.153
6, Bacterial diversity in rhizosphere soil from Antarctic vascular plants of Admiralty Bay, maritime Antarctica.
——(2010) ISME Journal 4(8): 989-1001.—— if 6.152
——(2010) FEMS Microbiology Letters 307(1): 80-6.—— if 6.152
8. Soil bacterial and fungal communities across a pH gradient in an arable soil. Â
——(2010) ISME Journal ePub:—— if 4.42
9, Examining the global distribution of dominant archaeal populations in soil. Â
—— (2010) ISME Journal ePub: —— if 4.42
10. Simultaneous assessment of soil microbial community structure and function through analysis of the meta-transcriptome .
—— (2008) PloS One 3:—— if 4.411
11, Characterization of trapped lignin-degrading microbes in tropical forest soil.
——(2011) PLoS One 6(4): e19306.—— if 4.411
12. Impact of biochar application to soil on the root-associated bacterial community structure of fully developed greenhouse pepper plants. Â
——(2011) Appl Environ Microbiol ePub:—— if 3.8
——(2011) Appl Eviron Microbiol ePub:—— if 3.8
——(2011) Appl Environ Microbiol ePub: —— if 3.8
15. Pyrosequencing-based assessment of bacterial community structure along different management types in german forest and grassland soils.
——(2011) PLoS One 6(2): e17000.—— if 3.8
16, Assessment of soil fungal communities using pyrosequencing. Â
——(2010) Journal of Microbiology 48(3): 284-9.—— if 3.8
——(2009) Environmental Microbiology ePub:—— if 3.778
18. Assessment of bias associated with incomplete extraction of microbial DNA from soil. Â
Feinstein LM, Sul WJ, Blackwood CB. (2009) Appl Environ Microbiol 75(16): 5428-33.—— if 3.778
19. Characterization of denitrification gene clusters of soil bacteria via a metagenomic approach. Â
——(2009) Applied Environmental Microbiology 75(2): 534-7—— if 3.778
- (2008) Soil Biology and Biochemistry 40: 2762-2770 - if 3.242
21, Shifts in microbial community structure along an ecological gradient of hypersaline soils and sediments. Â
——(2010) ISME Journal 4(6): 829-38.—— if 2.039
22. Metagenomic comparison of direct and indirect soil DNA extraction approaches.
——(2011) Microbiol Methods ePub: —— if 2.018
23, Horizon-specific bacterial community composition of German grassland soils as revealed by pyrosequencing-based analysis of 16S rRNA genes.
—— (2010) Appl Environ Microbiol 76(20): 6751-9.
references
[1] Li F D. Prosperous Areas of Current Soil Microbiology. Acta Pedologica Sinica , 1993 , 30(3) : 229-236.
[2] Xu GH , Li Z G. Microbial Ecology. Nanjing : Southeast University Press , 1991.
[3] Luo S M. Agroecology. Beijing : Chinese Agriculture Press , 2001. 72-96.
[4] Fromin N , Hamelin J , Tarnawski S , et al . Statistical analysis ofdenatinguring gel electrophoresis ( DGE ) finger printing patterns [ J ] .Environ Microbiol , 2002 , 4 (11) :634 - 643
[5] Ferris MJ, Muyzer G, Ward D M. Denaturing gradient gel electrophoresis profiles of 16S rRNA defined populations inhabiting a hotspring microbial mat community[J] . Appl Environ Microbiol , 1996 ,62 :340 - 346
[6] Zhang T, Fang HH P. Biotechnol Lett, 2000, 22: 399-405.
[7] Wagner M, Arnann R, Lemmer H, et al. Appl Environ Microbiol, 1993, 59: 1520~1525.
[8] Andreas F, Arthur W, Robert L, et al. FEMS Microbiol Ecol, 1999, 30: 137~145.
[9] Pyrosequencing enumerates and contrasts soil microbial diversity. Luiz FW Roesch, Roberta R Fulthorpe, Alberto Riva, George Casella, Alison KM Hadwin, Angela D Kent, Samira H Daroub, Flavio AO Camargo, William G Farmerie and Eric W Triplett (2008) The ISME Journal 1: 283–290
[10] Assessment of soil fungal communities using pyrosequencing. Lim YW, Kim BK, Kim C, Jung HS, Kim BS, Lee JH, Chun J. (2010) Journal of Microbiology 48(3): 284-9.
[11] Archaea predominate among ammonia-oxidizing prokaryotes in soils S. Leininger, T. Urich, M. Schloter, L. Schwark, J. Qi, GW Nicol, JI Prosser, SC Schuster, C. Schleper (2006) Nature 442: 806-809
[12] Simultaneous assessment of soil microbial community structure and function through analysis of the meta-transcriptome Urich T, Lanzén A, Qi J, Huson DH, Schleper C, Schuster SC. (2008) PloS One 3:
[13] A comprehensive survey of soil acidobacterial diversity using pyrosequencing and clone library analyses. Jones RT, Robeson MS, Lauber CL, Hamady M, Knight R, Fierer N. (2009) ISME Journal 3(4): 442-53
[14] Validation of heavy-water stable isotope probing for the characterization of rapidly responding soil bacteria. Aanderud ZT, Lennon JT. (2011) Appl Eviron Microbiol