EIGC Methods

The facility is CLIA certified (CLIA:11D1086150).  Our SOPs include DNA and RNA extractions from FFPE, fresh blood, frozen blood, PBMC, buffy coat, saliva, stool, tissue, cell pellet, and swabs (vaginal, rectal, buccal) samples.  We have the capacity to perform DNA and RNA extractions from other custom samples.  Read more about CLIA services.

We offer mutagenesis and subcloning services for your cloning needs.  Read more about custom cloning.

Library Sequencing

EIGC can provide library quantification services and sequencing services on our Illumina NextSeq 550 and MiSeq instruments.  EIGC can help you to outsource larger sequencing projects.

Whole genome sequencing

Whole genome sequencing is a comprehensive method for evaluating the entire genome of a sample.  Each sample requires around 90 Gb of sequencing.  The EIGC will help the investigator outsource the whole genome sequencing project, finding the best platform and pricing for their needs. 

Whole exome sequencing

Whole exome sequencing is a method to evaluate the protein-coding regions of the genome, allowing one to detect coding variants in a sample.  Whole exome sequencing may be a more cost-effective approach than whole genome sequencing for some scientific questions.  Each exome requires around 4-5 Gb of sequencing. The EIGC will help the investigator outsource the whole exome sequencing project, finding the best platform and pricing for their needs.

Single cell sequencing

Single cell sequencing is a technology that barcodes the DNA from individual cells with unique barcodes, allowing independent detection of genomes from each cell.  EIGC has the 10X Genomics Chromium Controller.  Single cell assays available with the 10X Genomics Chromium Controller are Single Cell RNA-Seq, Single Cell Immune Profiling, Single Cell ATACseq, and Multiome. See RNA-Seq (Single Cell) in the Transcriptomics section below for more information.

Targeted sequencing

Targeted sequencing allows Next Generation Sequencing of specific genes of interest.  EIGC works with investigators to design primer assays using the Fluidigm D3 Design tool, and we use the Fluidigm 48.48 Advanta Targeted Sequencing assays on the Access Array system to prepare libraries for sequencing.  Each LP 48.48 IFC allows NGS of up to 48 samples, using up to 4800 multiplexed assays.

Taqman genotyping and gene expression

Applied Biosystems’ TaqMan real-time PCR assays consist of target-specific primers and one or more probes optimized for specific types of applications, including gene expression, SNP genotyping, miRNA, and copy number variation analysis.  EIGC will use our ABI 7900HT instrument to perform RT-PCR for your project.

Microarray

Microarrays are designed to capture specific genomic information from a sample, such as the presence of specific genes in a pathway, structural variants, SNVs, CNVs, methylation sites, or custom sequences.  EIGC can provide Infinium MethylationEPIC and Infinium CytoSNP-850K v1.2 microarray services in-house.  The Infinium HD Methylation Assay Protocol Guide scales methylation profiling to thousands of CpG loci per sample, resulting in interrogation of over 850,000 methylation sites quantitatively across the genome at single-nucleotide resolution.  The CytoSNP-850K BeadChip uses an exon-centric design to target 3262 genes.  EIGC can help you to outsource other microarray services.  Read more about microarray.

Droplet digital PCR

Droplet Digital PCR (ddPCR) is a method for performing digital PCR that is based on water-oil emulsion droplet technology. Using BioRad’s QX200 instrumentation, a sample is fractionated into 20,000 droplets, and PCR amplification of the template molecules occurs in each individual droplet.  Fluorescent probes designed to detect specific DNA sequences are utilized to determine the presence of a DNA target of interest.  PCR-positive and PCR-negative droplets are counted to provide absolute quantification of target DNA in digital form.  Assays include absolute quantification, copy number variant analysis, rare mutation and sequence detection, gene expression, and detection of genome editing events.

NanoString nCounter Technology

NanoString’s nCounter FLEX Profiler allows analysis of tens to hundreds of targets from DNA, RNA, and protein samples.  The NanoString panels are available in pre-curated or custom panels.  EIGC will help the investigator to determine the best panels for a project and will perform the assay on the nCounter FLEX Profiler using extracted DNA, RNA, or lysate.  Read more about NanoString.

RNA-Seq (Bulk)

RNA-Seq determines the expression and sequence of RNA transcripts in a sample.  The EIGC can perform RNA-Seq or will help the investigator outsource the RNA-Seq project, finding the best platform and pricing for their needs.  Read more about bulk RNA-Seq.

RNA-Seq (Single Cell)

Single cell RNA-Seq determines the expression profile of individual cells within a sample.  EIGC uses the 10X Genomics Chromium Controller to prepare single cell sequencing libraries.  Sequencing of the prepared libraries can be performed in-house or outsourced, depending on sample size and coverage needs.  Read more about single cell RNAseq.

Amplicon (16S rRNA)

16S rRNA sequencing determines the microbial populations present in a sample using the 16S rRNA variable regions which distinguish phylogenetic classes of prokaryotes.  The EIGC currently supports sequencing two different 16S rRNA gene regions: V4 only and V3-V4.  V4 has a higher read pass rate, but lower ability to discriminate taxa in a sample. V3-V4 is a longer fragment resulting in a lower read pass rate but generates data with greater power to classify taxa and is the recommended region for new projects.  EIGC will prepare amplicon libraries from cell, tissue, fecal, or swab samples and sequence the libraries in-house.  Read more about 16S.

Shotgun metagenomics

Shotgun metagenomics sequencing provides a platform for sensitive detection of unculturable microorganisms and those which are less abundant in culture. This method comprehensively samples all genes in a bacterial sample in order to determine the taxonomy present in each sample.  EIGC will help the investigator to outsource shotgun metagenomics services.

ATAC-Seq

ATAC-Seq generates a global map of accessible chromatin on as few as 100 cells and up to 50,000 cells.  EIGC will prepare and sequence the ATAC-Seq libraries in-house.  Read more about ATAC-Seq.

ChIP-Seq and Cut&Run Library Prep

ChIP-Seq/Cut&Run provides a map of DNA sequences which interact with specific chromatin-associated proteins.  EIGC will provide library prep and sequencing services for your ChIP’ed samples.  (We do not perform the ChIP assay, but we can help you to outsource this service).

Reduced Representation Bisulfite Sequencing (RRBS)

RRBS maps CpG sites in the genome using restriction enzymes which cut at CG-containing DNA.  This service can be a cost-effective alternative to bisulfite sequencing of the whole genome.  Read more about RRBS.

Confirm knock-out and knock-in genome edits.

Spatial profiling reveals the transcriptome or proteome within an area defined on a slide.

10X Genomics Visium spatial profiling utilizes a special slide with a grid of capture areas, and the transcriptome is determined within each capture area.

Nanostring GeoMx spatial profiling allows collection of transcriptomic or proteomic information from specific groups of cells or tissue within a selected region of interest.  The regions can be as large as 660 x 785 um in size and may be circles, squares, or custom polygons.