We provide microbiome marker gene amplification and sequencing services, beginning from biological samples (feces, other human fluids, soil samples, etc.) or from extracted DNA. We offer library prep for multiple marker genes (bacterial 16S rRNA, fungal ITS1/ITS2, and eukaryotic 18S rRNA), using UMGC-developed methods, which in a recent report in Nature Biotechnology were shown to provide more quantitatively accurate and qualitatively complete measurement of mock community reference standards than other commonly used protocols. (Gohl et al (2016). Nat Biotechnol. 34:942-9).
In addition, we also offer library prep for the bacterial 16S rRNA V4 region and the eukaryotic ITS1 region using the Earth Microbiome Project protocol.
Marker-gene sequencing is typically carried out using the Illumina MiSeq platform, but deeper sequencing on the Illumina HiSeq for larger-scale projects, or for metagenomic and meta-transcriptomic methods is also available. For more information and to initiate a project, please contact Allison MacLean (firstname.lastname@example.org).
The UMGC is actively engaged in the development and optimization of microbiome profiling methods and is happy to discuss or collaborate on microbiome methods projects through our Research and Development group. Inquiries for these projects should be directed to Daryl Gohl, Ph.D. (email@example.com).
A “bead-beating” method that uses abrasive beads to break the cell walls of microbes in stool and other samples.
|MoBio PowerSoil for Stool, Soil, and Other Environmental Samples.|
|Yields depend on sample type.||1-95||sample||$24.65||$28.35|
|Yields depend on sample type.||96-any||sample||$12.94||$15.16|
The UMGC provides two workflows, a UMGC-developed dual-indexing (DI) approach, and the Earth Microbiome Project protocol.
This protocol uses a two step process in which the marker gene of interest is first amplified using a set of adapter-tailed primers, followed by addition of flow cell adapters and sample-specific indices by overlap extension. Our published protocol is described in (Gohl et al (2016). Systematic improvement of amplicon marker gene methods for increased accuracy in microbiome studies. Nat Biotechnol. 34:942-9).
The Earth Microbiome Project (EMP) protocol is available for the bacterial 16S rRNA V4 region, and the eukaryotic ITS1 region. This protocol uses a single round of PCR amplification during which flow cell adapters and sample-specific indices are added to the PCR primers to enable multiplexing of samples.
|16S/18S. UMGC Dual-indexing.|
|Price per sample. Variable regions available include: bacterial V1-V3, V3-V4, V3-V5, V4, V4-V6, V5-V6, and eukaryotic 18S (V9), ITS1, and ITS2. Standard turnaround is within 4-6 weeks of sample submission.||1-any||sample||$10.95||$12.65|
|Expedited (one-week project turnaround) 16S/18S. UMGC Dual-indexing.|
|We offer an expedited service which returns final sequencing results ≤ 7 calendar days from sample submission, including MiSeq sequencing. This service is only available on a per 96-well plate basis, and “as available” (assuming a MiSeq sequencer is “open” for the project). This rate must be paired with the Expedited MiSeq 300-bp Paired-end 2x300 PE) run.||1||up to 94 samples||$992.48||$1,146.77|
|16S/18S. UMGC Dual-indexing.|
|Earth Microbiome Project Protocol (V4 and ITS1).||Price per sample. V4 and ITS1 regions only.||sample||$8.49||$9.84|
The Illumina MiSeq is a “desktop” sequencer specialized for quick, lower-output runs on single-lane flow cells.
For paired-end lanes, we produce ≥ 8 million reads (Version 2) or ≥ 16 million reads (Version 3) in both forward and reverse directions.
With only a single lane to scan, the MiSeq has the quickest run per cycle of any Illumina instrument, and consequently, achieves the longest read length (paired-end 300 cycles). The MiSeq is therefore ideal for targeted sequencing of long amplicons, including microbial 16S/18S/ITS sequencing.
|MiSeq. Version 2 – Paired-end.|
|150-bp Paired-end (2x150 PE).||1-any||lane||$1,389.10||$1,686.89|
|250-bp Paired-end (2x250 PE).||1-any||lane||$1,518.39||$1,843.90|
|MiSeq. Version 3 – Paired-end.|
|300-bp Paired-end (2x300 PE).||1-any||lane||$1,968.50||$2,390.52|
|300-bp Paired-end (2x300 PE) Expedited.||1-any||lane||$3,210.19||$3,698.95|
For sequencing or amplicon marker gene samples on the HiSeq, output is ≥ 60 million reads in both forward and reverse directions. For shotgun metagenomic or transcriptomic samples, output is ≥ 120 million reads in both forward and reverse directions.
|HiSeq 2500 Rapid Run Mode – Paired-end.|
|150-bp Paired-end (2x150 PE).||1-any||lane||$3,085.46||$4,379.74|
|250-bp Paired-end (2x250 PE).||1-any||lane||$3,804.16||$5,399.92|
A microbiome project requires a multi-step bioinformatics analysis in order to generate high-quality results. We currently offer analysis of 16S and ITS data using our bioinformatics pipeline which implements the Qiime 1.9.1 analysis software. The analysis includes filtering raw fastq files for primer and adapter dimer sequences, removing contaminating host sequences and chimeric sequences, clustering sequences into OTUs using the Qiime open-reference OTU calling method with the greengenes 16S reference or UNITE ITS reference, and calculating alpha and beta diversity metrics. These analysis results are summarized in a report which includes a series of quality-control plots, interactive 3D PCA plots, taxonomy summary plots and data summary tables. Example Report.
These analysis services are provided free of charge to UMN groups by the Informatics Institute. Contact Trevor Gould (firstname.lastname@example.org) for more information. Analysis is offered to external customers by the UMGC and costs are detailed below. External customers should contact Trina Kuriger (email@example.com) for additional information. If your data analysis needs go beyond what is supplied by the analysis pipeline contact the Research Informatics Solutions group (firstname.lastname@example.org) at the Minnesota Supercomputing Institute for support.
|16s Analysis||1-any||MiSeq run (single assay/project)||Free||$288.31 + $3/sample up to 384 samples, $1/sample thereafter|