EIGC Methods
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.
| Assay | Detection Range | Size Range |
|---|---|---|
| Bioanalyzer DNA 1000 | 0.5-50 ng/µl | 25-1000 bp |
| Bioanalyzer DNA High Sensitivity | 5-500 pg/µl | 50-7000 bp |
| Bioanalyzer RNA Nano | 25-500 ng/µl (total RNA) 25-250 ng/µl (mRNA) | |
| Bioanalyzer RNA Pico | 50-5000 pg/µl (total RNA) 250-5000 pg/µl (mRNA) | |
| Quantit dsDNA PicoGreen | 250 pg/ml-1000 ng/ml | |
| Quantit dsDNA Broad Range | 2-1000 ng | |
| Quantit dsDNA High Sensitivity | 0.2-100 ng | |
| Qubit dsDNA Broad Range | 100 pg/µl -1000 ng/µl | |
| Qubit dsDNA High Sensitivity | 10 pg/µl -100 ng/µl | |
| Quantit RNA RiboGreen High-Range | 20 ng/ml-1 ug/ml | |
| Quantit RNA RiboGreen Low-Range | 1 ng/ml-50 ng/ml | |
| Fragment Analyzer HS NGS | 5 pg/uL-500 pg/µl (Fragment) 50-5000 pg/µl (Smear) | 100-6000 bp |
| Fragment Analyzer HS Large Fragment 50 kb | 5 pg/uL-600 pg/µl (Fragment) 50-5000 pg/µl (Smear) | 75-48,500 bp |
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 Standard Biotools D3 Design tool, and we use the Standard Biotools Targeted Sequencing assays on the Standard Biotools X9 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 help you to outsource microarray services.
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.
EIGC can help you to outsource other spatial workflows.