Illumina Whole Exome Sequencing

Illumina Whole Genome Sequencing Illumina Whole Exome Sequencing

Due to the large size and cost of sequencing the entire human genome, a widely used alternative is the more affordable option of Whole Exome Sequencing. Exome sequencing primarily targets only the protein coding regions of the genome. Representing < 2% of the human genome, this approach focuses limited resources, sequencing reads, and bioinformatic compute time to the most well-characterized regions of the genome and the areas of greatest interest.

Exome sequencing is, by default, a panel-based sequencing test. Several exome capture kits are available, and each kit differs in many areas, such as enrichment strategies, target regions, and length of probes. As a result, kit selection impacts coverage uniformity and capture efficiency on a region-by-region basis and is imperative to obtaining informative results.

SeqCenter uses both the Twist Exome 2.0 and the Twist Mitochondrial panels to span a wide breadth of targets and provide deep coverage of all nuclear and mitochondrial genes linked to known clinically relevant phenotypes. The Twist 2.0 panel demonstrates superior completeness of coverage, capturing a total of 19,810 genes (36.5Kbp), and the additional Mitochondrial Panel covers all 37 mitochondrial genes (16.6 kbp). In addition, this panel combination includes some noncoding regions that are known to carry pathogenic or likely pathogenic variants. Additional details on the Twist Exome 2.0 and Mitochondrial panels, as well as an example of our WES methods, are available here.

Sample Requirements:

  • Total volume of 30 µL or more
  • Eluted into water or dilute Tris/TE buffer
  • Sample concentration greater than:
    • 10 ng/µL measured by Qubit
    • 20 ng/µL measured by Nanodrop
  • Fragment length of at least 400bp

What You Will Receive:

  • Two fastq files (2 x 151bp) for each sample
  • Sequencing stats summary
  • Materials and methods summary
  • Data shared via Box folder within 2 weeks

Packages & Pricing

Sequencing Package
(Minimum Read Count Per Sample)		 Price*
50x Exome Coverage
Minimum of 2Gbp exome data.		 $320.00
100x Exome Coverage
Minimum of 4Gbp of exome data.		 $355.00
200x Exome Coverage
Minimum of 10Gbp of exome data.		 $400.00
*Orders >48 Samples receive 5% discount

What is the difference between whole exome sequencing and whole genome sequencing?

Whole genome sequencing allows you to examine the complete genetic information within your sample. This sequencing effort requires an extremely high amount of sequencing data to generate the required coverage necessary to perform informative analyses. Whole Exome Sequencing (WES) primarily targets only the protein coding regions of the genome.

While WGS will always be considered the “gold standard,” WES can be an attractive alternative due to its focused range. The targeted WES approach provides extremely high coverage for a fraction of the cost of WGS at the same depth. By focusing on protein coding features, WES also inherently narrows the analysis to regions of the DNA that are better characterized and more likely to be causative in nature. This also reduces overall output sizes which are nontrivial for human data. As a high-impact, low-cost human sequencing approach, WES can be a cost-effective option for studies where very high depth of coverage is necessary, such as in rare variant detection in oncological studies.

An often-cited advantage is the reduced time required for WES. However, with the introduction of Illumina’s NovaSeq X Plus and PacBio’s Revio, in combination with SeqCenter’s guaranteed turnaround time of two weeks for both services, this “advantage” diminishes. With the release of these larger volume platforms, WGS, rather than WES, could be considered the more efficient form of sequencing from a long-term perspective, as well. After producing WGS data of the full genome once, the original sequencing data can be re-analyzed in the future as new targets are discovered.

Additional Resources

Illumina Whole Genome Sequencing

Our Illumina Whole Genome Sequencing packages provide cost-effective short read sequencing using a tagmentation-based Illumina DNA library preparation. In addition to accommodating a wide range of sequencing size needs, this approach produces high-quality short reads suitable for a plethora of downstream analyses.

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Contact:
91 43rd Street, Ste. 250
Pittsburgh, PA 15201
(878) 227-4915

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Full Flowcell:

We offer full flowcell sequencing services for libraries that have been prepared outside of our lab. For each library, we will need to know the loading concentration, phiX spike-in, and run structure. Data is delivered as a complete run folder and can be demultiplexed upon request if a sample sheet is provided.

SARS-CoV-2 Sequencing:

Our SARS-CoV-2 Sequencing services include an assembly on the SARS-CoV-2 reference genome as a scaffold. These packages provide roughly 200Mbp of paired end reads (2×151 bp) as fastq files and the assembly in fasta format.

We require a minimum sample volume of 15 µL of extracted material (or 250µL for VTM samples) and recommend that only samples with Ct values below 35 are sent (because submissions are not measured for viral load before processing). We offer services for both purified RNA and samples in VTM media that require viral RNA extraction. Please see our SARS-CoV-2 Sequencing page for more detailed information.

Microbiome Sequencing:

Please see our Microbiome Sequencing page for package details, including distributables and sample requirements.

Nanopore Sequencing:

Our Oxford Nanopore sequencing packages provide reads equal to the length of input DNA, delivered as a fastq file.

For each sample, we require a minimum volume of 60 µL, and at least 40 ng/µL dsDNA quantified by Qubit or other dye-based method (100 ng/µL by Nanodrop). Variant calling add-on analysis requires an annotated reference file. DNA extraction services are available for only specific categories of microbes.

Please see our DNA Sequencing: Long Read Sequencing page for more detailed information about the package, variant calling analysis, and extraction requirements.

PacBio:

Our PacBio services deliver HiFi reads with an average read length of 8-12kb for standard offerings and up to 25kb for full flowcell runs. All package names are listed by the minimum number of >Q20 reads that will be delivered. Both assembly and annotation are included with all packages.

For each sample, a minimum volume of 60 µL and at least 35 ng/µL dsDNA (quantified by Qubit or other dye-based methods) are required. SeqCenter strongly recommends utilizing our HMW DNA Extraction service for best results; however, we will also accept samples that are shipped on dry ice to avoid DNA integrity issues caused by the shipping process.

Please see our PacBio Sequencing page for more detailed information about our services.

RNA Sequencing:

We offer both rRNA depletion RNA squencing and polyA-tail enrichment mRNA sequencing. Both provide stranded, paired end reads (2x 151bp) as fastq files.

For each sample, we require a minimum volume of 20µL and at least 50ng/µL RNA. We also recommend, but do not require, that samples be DNAse treated and have a RIN score greater than 6. We also offer Basic and Intermediate RNAseq analyses, which require an annotated reference sequence.

Please see our RNA Sequencing page for more detailed information about both our RNA sequencing and analysis packages.

Illumina Sequencing:

The Illumina sequencing packages produce paired end reads (2×151 bp) delivered as fastq files.

For each sample, we require a minimum volume of 30 µL and at least 10 ng/µL dsDNA quantified by Qubit or other dye-based method (20 ng/µL by Nanodrop). Variant calling add-on analysis requires an annotated reference file. DNA extraction services are available for only specific categories of microbes.

Please see our DNA Sequencing: Whole Genome Sequencing page for more detailed information about the package, variant calling analysis, and extraction requirements.

For additional information about variant calling and short read assemblies, please see our Bioinformatics page.

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