Whole Genome Resequencing

Concordance between homozygotes and heterozygotes as a function of coverage.

Reference Sequence

Once you have the reference sequence for an organism, you can perform comparative sequencing or resequencing to characterize the genetic diversity within the organism’s species or between closely related species.

Previously, the ability to generate sufficient depth of coverage for variant detection across an entire genome was limited by the throughput of existing platforms. This issue was compounded for larger organisms such as human and mouse. And while the ultra high throughput of next generation sequencing platforms held great promise for large scale resequencing experiments, initial technologies lacked the accuracy required for cost-effective variant detection.

Whole Genome Resequencing with the SOLiD™ System

The SOLiD™ System provides both the throughput and accuracy you need for cost effective variant detection. With demonstrated throughputs of 100 GB of mappable data and accuracy greater than 99.94%, the SOLiD™ System enables variant detection with much lower coverage than competitive platforms due to much lower false positive rates.

Identify Genome-Wide SNPs and Structural Variations in a Single Run
Using mate paired libraries enables you to detect structural variation such as insertion, deletions, inversions, and translocations in addition to SNPs. The SOLiD™ System supports a broad variety of mate paired insert sizes (0.6-6 KB), providing you with the flexibility to detect a variety of structural changes.

Read our application note on Mate-Paired Libraries Detect/Define Genetic Rearrangements (PDF).

Products

Experimental Workflow: Find the Products You Need for Every Step

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Step 3: Construct DNA Library

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Mapped mate-paried sequence tags.

You can prepare fragment, paired-end, and mate-paired libraries for whole genome sequencing. We recommend using mate-paired libraries for more accurate mapping.

Products for Step 3: Construct DNA Library

SOLiD™ Mate-Paired Library Oligos Kit: Contains a set of oligonucleotide adaptors and primers for constructing mate-paired libraries.

SOLiD™ Fragment Library Oligos Kit: Contains oligonucleotide adaptors and primers for fragment library construction.

SOLiD™ Long Mate-Paired Library Construction Kit w/o Purification: Provides a fully optimized, validated, complete set of reagents (minus the purification portion) for the construction of ten 2x35 bp or 2x50 bp libraries.

SOLiD™ Long Mate-Paired Library Construction Kit: This kit provides a fully optimized, validated, complete set of reagents for the construction of ten 2x35 bp or 2x50 bp libraries.

SOLiD™ 2x25 Mate-Paired Library Construction Kit: This kit provides a fully optimized, validated complete set of reagents of the construction of ten 2X25 bp libraries. In addition to this kit, customers will need to obtain the SOLiD™ Mate-Paired Library Oligo kit to generate mate-paired libraries.

SOLiD™ Fragment Library Construction Kit with SizeSelect™ Gels: This kit provides a fully optimized complete set of reagents, columns, and our new SizeSelect™ gels to generate 10 fragment libraries using the SOLiD™ System standard fragment library protocol.

View more reagents, kits and bench-top devices on invitrogen.com

SOLiD™ DNA Multiplexing Kits, 1-96 and Modules

The SOLiD™ Fragment Library Barcoding Kits enable users to pool fragment libraries prior to emulsion PCR and then conduct multiplexed sequencing analysis.

SOLiD™ Library TaqMan® Quantitation Kit

The SOLiD™ Library TaqMan® Quantitation Kit provides a complete qPCR solution to determine the appropriate amount of library to input into emulsion PCR for all library types.

E-Gel® SizeSelect™ Gels

Allows you to separate and recover DNA for short read fragment library construction in just 15 minutes.

For research use only. Not intended for any animal or human therapeutic or diagnostic use.

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User Guide: SOLiD™ 4 System Library Preparation Quick Reference Card (PN 4445674B)

Step 4: Amplify & Sequence

After you construct your libraries, amplify the DNA using standard emulsion PCR. Next, deposit the beads on a slide for sequencing on the SOLiD™ Analyzer

Products for Step 4: Amplify & Sequence

Sequencing

SOLiD™ 4 System: The Applied Biosystems SOLiD™ 4 System is a revolutionary genetic analysis platform that enables massively parallel sequencing of clonally-amplified DNA fragments linked to beads. The sequencing methodology is based on sequential ligation with dye-labeled oligonucleotides. The SOLiD™ 4 System enables researchers to obtain higher-quality genomes at lower cost without the purchase of a new instrument.

Fragment Library Sequencing Reagents for SOLiD™ 4 System, Main Kits: These kits contain sequencing master mixes in an easy-to-use strip tube format for ligation sequencing of fragment libraries on the SOLiD™ 4 System. Reagent mixing is not required, minimizing tube mix ups and providing a convenient automated method for sequencing reagent handling.

Instrument Buffers for SOLiD™ 4 System: This kit contains a set of buffers needed for ligation sequencing on the SOLiD™ 4 System. The kit is sufficient for 110 cycles. The product is compatible only with the SOLiD™ 4 System. Store components at ambient, 2 °C to 8 °C, and at -15 °C to -25 °C.

Workflow Analysis Reagents for SOLID™ 4 System: This kit is for accurate qualification and quantitation of templated beads prior to your sequencing run. Use this kit to perform a workflow analysis (WFA) run on the SOLiD™ 4 Analyzer.

Emulsion PCR (ePCR)

SOLiD™ EZ Bead™ System: The new SOLiD™ EZ Bead™ System simplifies next-generation sequencing workflows by providing an automated solution for reproducible templated bead preparation with less than 1 hour of hands-on time. The system is specifically designed with system modularity and quality control check points for greater experimental control. The SOLiD™ EZ Bead™ System streamlines the sequencing workflow through significant time savings and support for up to 96 multiplexed libraries. With automation, minimal costs, and seamless effort, the SOLiD™ EZ Bead™ System dramatically increases sequencing productivity to further accelerate scientific discovery.

SOLiD™ EZ Bead™ System Reagents and Accessory Kits: The new SOLiD™ EZ Bead™ System simplifies next-generation sequencing workflows by providing an automated solution for reproducible templated bead preparation with less than 1 hour of hands-on time. The system is specifically designed with system modularity and quality control check points for greater experimental control. SOLiD™ EZ Bead™ System streamlines the sequencing workflow through significant time savings and support for up to 96 multiplexed libraries.

SOLiD™ EZ Bead™ Emulsifier Oil Pack Kit

Bead Deposition

SOLiD™ Bead Deposition Kits: These kits contain components for depositing templated beads onto a slide for ligation sequencing on the SOLiD™ Analyzer. The kits contain reagents for performing a 3’ end modification and buffers. These kits are available in a 10 full-scale reaction size and a 20 mini-reaction size.

SOLiD™ Bead Enrichment Kits: These kits contain components to enrich templated beads amplified by emulsion PCR. The kit includes enrichment beads and reagents. These kits are available in a 10 full-scale reaction size and a 20 mini-reaction size.

SOLiD™ Slide Kits: These kits contain activated slides and storage buffer for ligation sequencing on the SOLiD™ Analyzer.

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Step 5: Analyze Data

Using the data analysis tools listed below:

Align reads to reference sequence
Identify genetic variation (SNPs, insertions, deletions, copy number and structural variation)
Translate to base space
Visualize annotated data

Products for Step 5: Analyze Data

Data Analysis Tools

The tools you need for each step in the whole genome resequencing workflow:


Data Analysis Step	Applied Biosystems Tools	Applied Biosystems Software	3rd-Party Software
1. Align reads to reference in color space	Resequencing Analysis Pipeline	SETS* SOLiD BioScope**	SHRiMP BFAST SOCS Geospiza Genomatix
2. Generate quality metrics and perform mate pair rescue	Resequencing Analysis Pipeline	SOLiD BioScope**
3. Generate sequencing and alignment statistics	Resequencing Analysis Pipeline	SETS*
4. Identify Polymorphisms	Resequencing Analysis Pipeline Small Indel Inversion Tool Large Indel CNV SNP Tool	SOLiD BioScope**	CNV-Seq Genomatix Partek
5. Translate color space to base space	GFF Conversion Tool Base QV Tool SAM Conversion Tool	SOLiD BioScope**
6. Visualize in context of annotation		SAB	Biotique Genomatix GenomeQuest NextGENe Partek UCSC Browser
7. Convert to SRF for publishing	SRF Conversion Tools

*on-instrument **off-instrument

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Publications & Literature
Application Notes
- Whole Genome Sequencing of E. coli
- Application Note: SOLiD™ System Mate-Paired Libraries Detect and Define Large Genetic Rearrangements
Publications
- The complete genome sequence of Escherichia coli DH10B: insights into the biology of a laboratory workhorse. Publication: J Bacteriol. 2008 Apr;190(7):2597-606. Epub 2008 Feb 1. Authors: Durfee, T., et al.
- Whole-genome resequencing reveals loci under selection during chicken domestication. Publication: Nature 464, 587-591 25 March 2010 Authors: Rubin C., et al.
- Whole-genome sequencing in a patient with Charcot-Marie-Tooth neuropathy. Publication: N Eng J Med 10 March 2010 Authors: Lupski J., et al.
- Development of personalized tumor biomarkers using massively parallel sequencing. Publication: Science Translational Medicine, 24 February 2010 Authors: Leary R., et al.
- U87MG Decoded: The Genomic Sequence of a Cytogenetically Aberrant Human Cancer Cell Line. Publication: PLoS Genetics, January 2010, volume 6, issue 1 Authors: Clark, et al.
- Maternal Plasma DNA Analysis with Massively Parallel Sequencing by Ligation for Noninvasive; Prenatal Diagnosis of Trisomy 21 Publication: Clinical Chemistry 56:3 (2010) Authors: Chiu, et al.
- Complete Khoisan and Bantu genomes from southern Africa. Publication: Nature, Vol 463, 18 February 2010 Authors: Schuster et al.
- Whole-Exome Sequencing Identifies FAM20A Mutations as a Cause of Amelogenesis Imperfecta and Gingival Hyperplasia Syndrome Publication: American Journal Human Genetics, (2011) Authors: O'Sullivan J. et al.
- Comparison of Sequence Reads Obtained from Three Next-Generation Sequencing Platforms Publication: PLoS ONE (2011) Authors: Suzuki S. et al.
- Extensive genomic and transcriptional diversity identified through massively parallel DNA and RNA sequencing of eighteen Korean individuals Publication: Nature Genetics (2011) Authors: Ju. et al.
- Global analysis of disease-related DNA sequence variation in 10 healthy individuals: Implications for whole genome-based clinical diagnostics Publication: Genetics in Medicine (2011) Authors: Moore. et al.
- Demographic history and rare allele sharing among human populations Publication: PNAS 2011 Authors: Gravel S et al

For Research Use Only. Not intended for any animal or human therapeutic or diagnostic use.

Whole Genome Resequencing

Whole Genome Resequencing with the SOLiD™ System