High Throughput NGS
Intended for full-length sequencing of clonal circular plasmid DNA or PCR product above 0.5kb in length, and whole bacterial genome.
This service is performed using state-of-the-art Illumina® NextSeq NGS platform, and includes the following components:
- Construct an NGS library, including linearization of circular DNA and/or template amplification, with QC procedure at each step;
- Sequence the library primer-free with comprehensive coverage at high depth and accuracy;
- Align the reads against a reference sequence, or assemble the reads into a larger contig
In most cases, sequencing data and analysis results are delivered within 2 weeks of sample receipt.
<span data-metadata="">Service Description
- We receive your sample and notify you of sample receipt.
- Upon request, sample DNA template is amplified and QC’ed by Qubit fluorometry.
- Sample DNA is used to construct NGS library, and library is QC’ed by 2100 Bioanalyzer electrophoresis.
- NGS library pool is size selected and QC’ed by Qubit fluorometry and 2100 Bioanalyzer electrophoresis.
- Compatible NGS libraries are sequenced by Illumina NextSeq platform.
Bioinformatics Analysis
Standard types of bioinformatics analysis are included with High Throughput NGS
Map to Reference
Sequenced NGS reads are aligned to the reference sequence to detect genetic variations (SNVs, InDels, CNVs) and generate a consensus sequence.
Application: Confirming sample sequence and detecting sequence deviations
Deliverables: Full project report; NGS sequencing QC report; FASTA-format consensus sequence of sample reads; variant report.
De Novo Assembly
Sequenced NGS reads are constructed to a larger, scaffolded set of contigs, representative of the original sequence, without any reference.
Application: Obtaining sample sequence without prior knowledge.
Deliverables: Full project report; NGS sequencing QC report; FASTA-format best contig assembly from sample reads.
Note: De novo assembly is not recommended for episome analysis with bacterial culture or colony sample types, due to significant bacterial gDNA contamination. For bacterial genomes, de novo assembly may be incomplete and contain multiple shorter contigs.
Custom Analysis
More complex bioinformatics analysis for various research purposes, such as comparative genomic and transcriptomic analyses, methylation analysis, small RNA analysis, and cancer NGS data, can be arranged on a project-specific basis.
High Throughput NGS vs Sanger Sequencing
Unlike traditional Sanger sequencing, which relies on primers to evaluate a specific target region, High Throughput NGS service sequences the entire episome or PCR product molecule with high coverage depth and accuracy
- High Throughput NGS yields reads with quality of Q30 or higher (>99.9% accuracy)
- High Throughput NGS yields coverage of greater than 500X average depth over entire episome sequence
- High Throughput NGS is able to reveal base deviations and small indels, not detectable with Sanger, at higher sensitivity and lower limit of detection
- High Throughput NGS is more cost-effective for medium to large targets longer than 750 bp
- High Throughput NGS is the practical solution for bacterial whole genome sequencing (WGS)