454 Sequencing

Roche

Achieve More

Instrument & Workflow

A new scale of sequencing to meet your lab’s needs

The GS Junior System brings the power of 454 Sequencing technology directly to your laboratory bench top. Benefit from the same proven long-read sequencing performance as the Genome Sequencer FLX System, scaled to suit the needs of individual labs. Quickly proceed from DNA to data to discovery with a simple sample preparation workflow, overnight sequencing and data processing, and a dedicated suite of data analysis software.

System Performance
Throughput 35 million high-quality, filtered bases per run*
Run Time 10 hours sequencing
2 hours data processing
Avg. Read Length 400 bases*
Accuracy Q20 read length of 400 bases
(99% accuracy at 400 bases)
Reads per Run 100,000 shotgun, 70,000 amplicon
Sample Input gDNA, amplicons, cDNA, or BACs depending on the application
Physical Dimensions 40 cm wide x 60 cm deep x 40 cm high (the size of a laser printer)
Weight = 55 lbs.
Computing Linux-based OS on HP desktop computer included.
All software is point-and-click.
*Typical results. Average read length and number of reads depend on specific sample and genomic characteristics

 

Workflow

The GS Junior System offers an end-to-end sequencing solution from sample preparation and sequence generation through data analysis. Robust protocols with minimal handling steps make the workflow ideally suited for individual labs.

  • Produce libraries in less than half a day with easy-to-follow sample preparation protocols
  • Use only general laboratory equipment without the need to purchase tons of additional supplies
  • Perform overnight sequencing and data processing with a quick 10-hour instrument run time
  • Go from sequence data to publishable result with straightforward tools for de novo assembly, mapping and amplicon variant analysis

How It Works

1. Sample Input and Fragmentation
The GS Junior System supports the sequencing of samples from a wide variety of starting materials including genomic DNA, PCR products, BACs, and cDNA. Samples such as genomic DNA and BACs are randomly fragmented into small, 300- to 800-basepair pieces. For smaller samples, such as small non-coding RNA or PCR amplicons, fragmentation is not required. Instead, PCR products amplified using Genome Sequencer fusion primers may be immobilized onto DNA capture beads and clonally amplified as shown below under "One Fragment = One Bead".

2. Library Preparation
Using a series of standard molecular biology techniques, short DNA adaptors are added to each library fragment. These adaptors are then used in subsequent quantification, amplification, and sequencing steps.

3. One Fragment = One Bead
The single-stranded DNA library is immobilized onto specifically designed DNA Capture Beads. Each bead carries a unique single-stranded DNA library fragment. The bead-bound library is emulsified with amplification reagents in a water-in-oil mixture resulting in microreactors containing just one bead with one unique sample-library fragment.

4. emPCR (Emulsion PCR) Amplification
Each unique sample library fragment is clonally amplified within its own microreactor, excluding competing or contaminating sequences. Amplification of the entire fragment collection is carried out in parallel; for each fragment, this produces several million copies of the original fragment per bead. Subsequently, the emulsions are broken to facilitate collection of the amplified fragments bound to their specific beads.
 
5. One Bead = One Read
The clonally amplified fragments are enriched and loaded onto a PicoTiterPlate device for sequencing. The diameter of the PicoTiterPlate wells allows for only one bead per well. After addition of sequencing enzymes and reagents, the fluidics subsystem of the Genome Sequencer System serially flows nucleotides in a fixed order (i.e. first T, then A, and so on) across the hundreds of thousands of wells containing one bead each. Addition of one (or more) nucleotide(s) complementary to the template strand results in a chemiluminescent signal recorded by the CCD camera of the Genome Sequencer System. The intensity of the resulting signal is proportional to the number of bases incorporated.

6. Data Analysis
The combination of signal intensity and positional information generated across the PicoTiterPlate device allows the software to determine the sequence of 100,000 individual reads per 10-hour instrument run simultaneously. For sequencing data analysis, three different bioinformatics tools are supplied that readily support the following applications: de novo genome assembly up to 3 Gb; resequencing/mapping genomes of any size; and amplicon variant detection by comparison with a known reference sequence.