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Microbiome Multi-omic Services

Metagenomics analysis starts with DNA extraction. Then, amplicon sequencing or whole genome sequencing (WGS) is performed. Amplicon sequencing is for analysis of a specific targeted gene (16S,18S or ITS). Shotgun metagenomics amplifies all the DNA of a mixed community which enables higher resolution and functional data analysis.

Figure 1.Metagenomics analysis starts with DNA extraction. Then, amplicon sequencing or whole genome sequencing (WGS) is performed. Amplicon sequencing is for analysis of a specific targeted gene (16S,18S or ITS). Shotgun metagenomics amplifies all the DNA of a mixed community which enables higher resolution and functional data analysis.

We have augmented the capabilities of our Sigma-Aldrich® portfolio of microbiome research products with multi-omics services to enhance analysis of diverse biological and environmental samples, and to provide robust and comprehensive detection and analytical services—from sample preparation to statistical reporting.


HOW MULTI-OMICS ANALYSIS SERVICES ACCELERATE MICROBIOME RESEARCH

Our microbiome services pipeline empowers microbial community analysis with:

  • A comprehensive collection of multi-omics services, in one place: our capabilities include both metagenomics and metabolomics services.
  • Services for the complete workflow: from low-bias DNA extraction to bioinformatic results, at the click of a button.
  • Identification and quantification of samples from varied sources: fecal, saliva, skin, mucosa, swab, soil, and many more.
  • Resolution at the species level with accurate, credible results: Our defined protocols include internal standards to assure minimal workflow bias.
  • The M-CAMPTM system: An easy-to-use, cloud-based, interactive bioinformatic platform. Try it now, at no cost.
  • Partnership with our microbiome research scientists: our R&D scientists consult with and assist our customers to ensure meaningful results from your microbiome studies

Our Low-Bias Metagenomic Services

Our substantial microbiome research expertise has enabled us to develop highly accurate metagenomics sequencing services for characterization of the gut, skin, and many more important microbial environments. Our extraction protocol, library preparation, QC, and sequencing process are all controlled with our in-house-developed microbial standards, in accordance with recommended best practices1 that assure low-bias results. Combining Illumina sequencing platforms with our M-CAMPTM bioinformatics software will enhance the power of your data. This data synergy increases taxonomic resolution by enabling species, sub-species, and strain level profiling of the composition of your sample, even with reduced read depth.

Our metagenomic services include amplicon metagenomics sequencing, to enable microbial profiling by bacterial / fungal gene markers (16S, 18S or ITS) by this rapid and cost-effective method.

WHAT ARE THE BENEFITS OF THE AMPLICON METAGENOMIC SEQUENCING METHOD?

Features of the amplicon sequencing of phylogenetic markers include

  • Low cost
  • Resistance to host DNA contamination
  • Allows identification of extremely low abundance microbes

16S rRNA metagenomic sequencing for bacterial identification

The 16S rRNA gene is highly conserved in bacteria, and serves as a common bacterial gene marker. The 16S rRNA metagenomic sequencing method is used to identify, classify and quantify the bacteria within a complex sample. The usage of PCR primers, targeted for specific conserved regions of 16S rRNA gene, enables the sequencing of neighboring variable regions. These regions contain taxonomic information which enable the identification of different bacteria present within a sample.

5R-Plex ultrasensitive 16S library preparation technology overcomes low biomass or degraded DNA challenges

Our experts developed highly sensitive, low-bias 16S amplicon metagenomic sequencing. Use our exclusive 5R-Plex Ultra-sensitive 16S library preparation technology. The 5R-Plex system is based on the SMURF (Short Multiple Regions Framework) technology developed to facilitate cancer microbiome studies. The 5R-Plex approach is based on amplifying 5 short regions (~200bp) along the 16S rRNA gene, followed by the usage of our exclusive algorithm to provide a larger coverage of the 16S rRNA gene. SMURF-based 5R-Plex analysis allows better resolution during microbial community profiling - even in extreme conditions like low bacterial biomass and fragmented DNA2,3,4.

18S rRNA / ITS metagenomic sequencing:

The 18S rRNA gene is highly conserved in eukaryotic cells. ITS (Internal Transcribed Spacer) regions are spacer DNA located between the small and large ribosomal subunit 18S genes. These markers are therefore useful for identifying, classifying, and quantifying fungi. Although ITS1&2 are popular fungal gene markers, the 18S rRNA gene has been shown to outperform these markers for taxonomy composition and resolution5.

To facilitate gut, oral, skin and vaginal human mycobiome studies, we’ve developed a highly sensitive protocol which enables the detection of very low abundance fungi (0.001 - 0.1% of the microbial community) with minimal host detection.

BENEFITS OF THE SHOTGUN METAGENOMIC SEQUENCING METHOD, OR WHOLE GENOME SEQUENCING (WGS)

Whole genome sequencing (WGS), also known as shotgun metagenomic sequencing, is a method that breaks the genome into a collection of small DNA fragments that are then individually sequenced. Attributes of WGS include:

  • Provides a detailed understanding regarding the composition of microorganisms in a sample
  • WGS enables sampling the entire gene pool of all the organisms that are present in a complex sample
  • High taxonomy resolution to the strain level, unlike amplicon-based methods including 16S rRNA or ITS sequencing
  • WGS can be used to characterize microorganisms from any taxa, and increases resolution by providing information about the strain level variation.
  • Identification of all members of the microbial community, as WGS detects even low abundance microbes
  • Identification of the entire genome using WGS reveals the potential functional profile of the microbiome of interest, elucidating enriched genes and pathways.
  • Low primer & PCR bias: our experts have developed PCR-free protocols aligned with best practices for library construction6

PCR-FREE ANALYSIS: LOW-BIAS SHOTGUN METAGENOMICS

During shotgun library preparation, amplification is typically performed for sequencing, introducing technical bias to the microbiome results. For samples with high DNA abundance, we offer PCR-free shotgun analysis to significantly reduce bias.

Shotgun Metagenomics for Low-Input DNA Samples

Many biological samples end up with very low DNA input. Our shotgun protocol is suitable for samples as low as 0.5ng/ml DNA input, with no bias.

THE M-CAMP™ PLATFORM: BIOINFORMATIC ANALYSIS SUITABLE FOR COMPLEX MICROBIOME DATA

The M-CAMP™ (Microbiome Computational Analysis for Multi-omic Profiling) platform is an intuitive, cloud-based software tool that does not require annual licenses or complicated downloads. This user-friendly platform provides:

  • A choice of standardized and updated sequencing databases, including a proprietary 16S database (V3-V4 region)
  • Key criteria selection for QC processing of sequencing data
  • Statistical and graphical analyses of microbiome communities
  • Publication-ready, downloadable graphics and reports: alpha diversity boxplots, PCoA (principal coordinates analysis), cladogram, LEfSe biomarkers, and taxonomy abundance heatmaps, all at your fingertips
  • Tools for taxonomic classification of bacteria within a single sample or for inter-sample comparative analysis

The M-CAMPTM 16S-seq taxonomic classification algorithm provides species-level classification using a novel methodology and a curated database built by combining reference sequences from multiple sources from which the user can choose, including Refseq prokaryote genomes, the Refseq Targeted Loci 16S ribosomal RNA project, SILVA release 138 and a proprietary Sigma-Aldrich® 16S database. Compared with current popular public classification algorithms, our algorithm provides the most accurate species-level classification of 16S rRNA sequencing data. For your convenience, we include the option to upload sample metadata. Try the M-CAMP™ system now, free of charge.

Our experts can assist in your upstream clinical study design with microbiome biological sample collection, preparation, statistical analysis, and downstream reporting over the course of your service project. Click here to request our services.


References

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Nearing JT, Comeau AM, Langille MGI. 2021. Identifying biases and their potential solutions in human microbiome studies. Microbiome. 9(1): http://dx.doi.org/10.1186/s40168-021-01059-0
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Fuks G, Elgart M, Amir A, Zeisel A, Turnbaugh PJ, Soen Y, Shental N. 2018. Combining 16S rRNA gene variable regions enables high-resolution microbial community profiling. Microbiome. 6(1): http://dx.doi.org/10.1186/s40168-017-0396-x
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Amir A, Zeisel A, Zuk O, Elgart M, Stern S, Shamir O, Turnbaugh PJ, Soen Y, Shental N. 2013. High-resolution microbial community reconstruction by integrating short reads from multiple 16S rRNA regions. 41(22):e205-e205. http://dx.doi.org/10.1093/nar/gkt1070
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Nejman D, Livyatan I, Fuks G, Gavert N, Zwang Y, Geller LT, Rotter-Maskowitz A, Weiser R, Mallel G, Gigi E, et al. 2020. The human tumor microbiome is composed of tumor type?specific intracellular bacteria. Science. 368(6494):973-980. http://dx.doi.org/10.1126/science.aay9189
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Frau A, Kenny JG, Lenzi L, Campbell BJ, Ijaz UZ, Duckworth CA, Burkitt MD, Hall N, Anson J, Darby AC, et al. 2019. DNA extraction and amplicon production strategies deeply inf?luence the outcome of gut mycobiome studies. Sci Rep. 9(1): http://dx.doi.org/10.1038/s41598-019-44974-x
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Tourlousse DM, Narita K, Miura T, Sakamoto M, Ohashi A, Shiina K, Matsuda M, Miura D, Shimamura M, Ohyama Y, et al. 2021. Validation and standardization of DNA extraction and library construction methods for metagenomics-based human fecal microbiome measurements. Microbiome. 9(1): http://dx.doi.org/10.1186/s40168-021-01048-3