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Please, download and complete the Contact Information Form and the specific form for the requested service, and bring them along with your samples.

High Throughput Sequencing

16S rRNA Amplicon Sequencing
Sequencing of targeted variable regions (V3-V4) in the 16S ribosomal gene allows to determine the bacterial components of microbial complex communities. Data provided include relative abundance of taxa at the genus/species-level, alpha and beta diversity calculations and basic analysis of significantly over or underrepresented groups.
Whole Genome Shotgun (WGS) Sequencing
WGS sequencing generates high depth sequence data (>10M sequences per sample) of entire microbial communities for higher resolution taxonomic characterization compared to 16S rRNA sequencing, providing species/strain-level information and community functional potential (enzymatic pathways and gene families). WGS can also be applied to isolated microbial strains to generate genomic sequencing data.
RNA Sequencing
Total community meta transcriptomics generates sequencing data from genes being actively expressed across community members. RNA sequencing can also be used to target host-specific factors, or to characterize changes in expression of individual bacterial strains in culture.

Shallow WGS Sequencing
Whole Genome Shotgun (WGS) sequencing performed at lower sequencing depth (>1M reads per sample) is a cost-effective method to acquire species/strain-level taxonomic characterization of the more abundant members of complex microbial communities. By sequencing at a lower depth, the cost of sequencing is reduced, however this will reduce the ability to accurately assess the identity or potential function of low-abundance community members. This approach is offered as a lower-cost alternative to WGS sequencing while providing higher taxonomic depth than standard 16S rRNA amplicon sequencing.

Sequencing Request Form

Quantitative (q)-PCR

Standard qPCR
The method is based on the conserved 16S rRNA gene present in all bacteria (a comprehensive list of 16S rRNA sequences from gut bacteria have been deposited and categorized in the Ribosomal Database Project II ). By calculating the number of conserved 16S rRNA genes, qPCR can indirectly indicate the number of bacteria present in a given sample. Additionally, appropriate design of qPCR oligonucleotide probes (primers) that can identify the sequence of the 16S gene variable regions allows for the amplification of genes from a specific bacterium of interest whilst excluding all other bacteria, making qPCR a useful tool for quantifying specific bacteria in a complex microbial community.
Fluidigm Access array targeted high-throughput qPCR

The BioMark™ Systems paired with the Fluidigm Dynamic Arrays are an efficient solution for large-scale, real-time qPCR. The nanofluidic chips contain fluidic networks that automatically combine sets of samples with sets of assays. This innovative solution for real-time qPCR provides reaction densities far beyond what is possible with microtiter plates and significantly reduces the number of liquid-handling steps and the volume per reaction.
Digital PCR
This technology expands the application boundaries of traditional real-time PCR. In contrast to real-time PCR this platform enables absolute quantification without the use of a standard curve and reference sample allowing a detection of individual DNA molecules with higher accuracy for a variety of applications.

The chip used in this system contains 20,000 individual wells and works by partitioning a standard PCR reaction into thousands of individual PCR reactions. A portion of these partitions contains the target molecule, while other partitions do not, leading to positive and negative reactions. Following amplification, the fraction of negative reactions is used to quantify the number of target molecules in the sample, all without reference to standards or controls.

We offer digital PCR service using QuantStudio 3D™ Digital PCR system from ThermoFisher Scientific. This is a chip based digital PCR platform. In contrast with droplet digital PCR the reaction is taking place in a well of the chip and not in a droplet, with fewer pipetting steps, which decreases the likelihood of cross-contamination.

Applications:

  1. Absolute quantification of DNA molecules
  2. Pathogen detection and load determination
  3. GMO detection and contamination assessment
  4. Library quantification for next-generation sequencing
  5. Characterization of low-fold changes in mRNA and miRNA expression
  6. Rare cancer mutation quantification
  7. CNV detection

Advantages:

  1. Digital PCR does not rely on Ct values to quantify copy number, thus, PCR efficiency differences among biological samples are much less likely to affect quantitative results.
  2. Digital PCR does not rely on standard curve for absolute quantification.
  3. Digital PCR is much less sensitive to PCR inhibitors present in crude samples.
qPCR Analysis Service Request Form

Consulting and Research Support

Including:

  • Sample genomic DNA isolation
  • Plasmid and virus nucleic acids isolation
  • RNA isolation
  • Strain typing
  • Optimization of bacterial culture conditions
  • High-throughput liquid handling (PCR reaction set up, sample pooling, picogreen quantification of nucleic acids)

AMC Culture Collection

The understanding of host-associated and free-living microbial communities’ structure and functionality has greatly advanced via high-throughput sequencing. As the field moves from association studies to mechanistic and interventional studies, the availability of authenticated, reliable biological material and associated information has become crucial.

In partnership with the Center for Gastrointestinal Biology and Disease (CGIBD) Advanced Analytics Core (AAC) and the Carolina Donor Services (CDS), the federally designated organ procurement organization serving 7.2 million people in 77 counties of North Carolina and Virginia, the Microbiome Core has isolated, characterized and preserved over 500 human isolates and reference probiotic bacteria of medical and scientific relevance.

The AMC Culture Collection adds to our current services of microbiome analysis to provide well-characterized, active and viable strains for microbiome studies that demand specific bacterial groups and/or functionality. Contact us at microbiome@med.unc.edu for more information or a consultation.

Take a look!

New Services:

  • Strain/consortium request: This service provides well-characterized human isolates and reference probiotic bacteria for research. Lyophilized, physiologically active or glycerol stocks can be provided as requested.
  • Strain submission for storage and preservation: Researchers can submit relevant strains for long term storage and preservation. Service can include strain characterization (antibiotic resistance, carbohydrate utilization and genome sequencing).
  • Activation and culturing of strains: The service includes the activation of bacterial cultures under physiologically appropriate conditions. The Core will provide cultures in liquid or solid media as requested.