Whole Genome Sequencing
Get comprehensive genome coverage and explore the full range of genetic variation using long, high-quality reads from the GS FLX and GS Junior Systems. Perform straightforward de novo assembly to decode previously uncharacterized genomes, or resequence organisms with an available reference. Use shotgun reads alone or in combination with multi-span paired end reads (3 kb, 8 kb, 20 kb) to tackle any size genome from bacteria to large, complex plants and animals.
- Quickly generate high-quality reference genome assemblies by assembling sequence reads de novo
- Accurately annotate genes in whole genome assemblies
- Compare sequence reads with a reference for variant calling of SNPs, insertions, deletions, inversions, and structural rearrangements of any size
- Carry out genome-level comparative analysis, such as synteny and identification of horizontal gene transfers and orthologs
- Near-finished de novo assemblies of microbial genomes in days — Use the GS De Novo Assembler software to obtain a high-quality draft assembly in about 15 minutes, with minimal prior experience in bioinformatics.
- The power to sequence large, complex genomes — Long reads sequence through repeat regions directly, closing gaps in contigs and minimizing expensive follow-up experiments. Combine shotgun reads with 3 kb, 8 kb, and 20 kb multi-span paired end libraries to orient genomic fragments and identify structural variations.
- New! GS FLX+ System read lengths up to 1 kb — Extra-long sequencing reads improve assembly quality by spanning even larger repeat regions and resolving breakpoints into contiguous sequence, resulting in assemblies with fewer, longer contigs and scaffolds.
- Perform hybrid assemblies — Combine long GS FLX+ shotgun reads, multi-span paired end reads and short read data for reduced project costs. The GS De Novo Assembler and GS Reference Mapper support input of FASTQ data files for simple co-assembly with short read data and data from sequence read archives.
Single Scaffold Assemblies of Microbial Genomes
| Microbial Genome Assembly | ||||
|---|---|---|---|---|
| Organism | S. pneumoniae | E. coli | T. thermophilus | C. jejuni |
| Number of chromosomes | 1 | 1 | 2 | 1 |
| Large Scaffolds | 1 | 1 | 2 | 1 |
| Genome Size | 2.2 Mb | 4.6 Mb | 2.1 Mb | 1.6 Mb |
| N50 Scaffold Size | 2.2 Mb | 4.6 Mb | 1.9 Mb | 1.6 Mb |
| N50 Contig Size | 26.1 Kb | 57.1 Kb | 10.5 Kb | 153.8 Kb |
| Genome Coverage | 99.6% | 100% | 100% | 99.3% |
| Oversampling | 25x | 15x | 33x | 33x |
| Number of Runs | 1/4 | 1/4 | 1/4 | 1/4 |
Table: Assembly of four microbial genomes into a single, large scaffold per chromosome using data generated on a single GS FLX Titanium run (4-region gasket). Assemblies were generated using one 8 Kb paired end library per genome.
454 Sequencing Systems offer a variety of approaches for your whole genome sequencing projects. Use shotgun reads alone or in combination with paired end reads to generate accurate draft assemblies of any organism.
Highly versatile, long shotgun reads open the full spectrum of genomic variation to analysis. Whether the targets are SNPs, insertions or deletions, novel genes or genomic regions, long shotgun reads enable you to investigate the entire genome more completely. Use shotgun methods to sequence purified gDNA from a microbe, plant, animal, fungus, virus, BACs or any other DNA source. Assemble shotgun reads into contigs using the GS De Novo Assembler or map them against an available reference sequence using the GS Reference Mapper.
The GS Junior System generates shotgun sequencing reads ~400 bp in length. The GS FLX+ System offers sequencing read lengths up to 1 kb using the GS FLX Titanium Sequencing Kit XL+.
Figure: Assembly of shotgun reads into contigs using the GS De Novo Assembler Software
The combination of multi-span paired end (3 kb, 8 kb, 20 kb) reads and shotgun reads offers a powerful solution for end-to-end de novo sequencing and high quality draft assembly. Generate assemblies with longer contigs and fewer, larger scaffolds – saving time and resources for post-sequencing finishing. Paired end reads provide the largest benefit with complex genome assemblies such as plants and animals, when multiple repeat regions longer than the typical read length are present, or when large structural rearrangements need to be accurately identified and located.
Use the GS FLX or GS Junior System to perform paired end sequencing and benefit from:
- Long tags - Paired end reads with long tags, averaging 140-200+ bases, can be aligned uniquely with high confidence
- Long spans - Long 20 kb paired end reads span most repeat regions in nearly any size genome
- Span variety - Broad selection of insert lengths (3 kb, 8 kb, 20 kb) enables optimization of project design according to the unique characteristics of any genome
Overview of the paired end protocol for use on the GS FLX or GS Junior Systems. Genomic DNA is sheared into 20 kb, 8 kb or 3 kb fragments. Adaptors are added to the end of each fragment, facilitating circularization. The circularized DNA is fragmented and fragments containing the added adaptors are isolated for sequencing, and generate true paired end reads with two end tags averaging over 140 bp and separated by 20 kb, 8kb or 3kb.
GS De Novo Assembler Software: Produce high-quality draft assemblies for microbial genomes in less than 15 minutes, and in less than 24 hours for larger genomes.
GS Reference Mapper Software: Rapidly and accurately align reads to any reference genome and explore the full range of genomic variations: SNPs, insertions, deletions and structural variations.
- The genome sequence of the leaf-cutter ant Atta cephalotes reveals insights into its obligate symbiotic lifestyle. Suen et al. (2011) PLoS Genetics 7(2):e1002007.
- The genome of the Theobroma cacao. Argout X et al. (2011) Nature Genetics 43(2):101-8.
- Complete Khoisan and Bantu genomes from southern Africa. Schuster SC et al. (2010) Nature 463(7283): 943-7.
- The genome of the domesticated apple (Malus × domestica Borkh.) Velasco R et al. (2010) Nature Genetics 42(10): 833-9.
- Rapid Identification of Genetic Modifications in Bacillus anthracis Using Whole Genome Draft Sequences Generated by 454 Pyrosequencing. Chen PE et al. (2010) PLoS One 5(8): e12397.
Search the complete list of 454 Sequencing publications here.
Literature
- De Novo Sequencing of Small Genomes Flyer
- De Novo Sequencing of Complex Genomes Flyer
- 454 Sequencing Systems in Pathogen Detection (Flyer)
Video