Sequencing

Sequencing

Overview

The primary goal of the sequencing core facility at LJI is to apply the latest sequencing technology combined with the Institute’s expansive bioinformatics capabilities to enrich research at the Institute and beyond. High-throughput sequencing is an extremely fast developing technology that has profoundly altered our understanding of biology, human diversity, and disease.

Instrumentation

Hiseq 2500

The Flagship of the Illumina family, the Hiseq 2500 uses Illumina’s Sequencing by Synthesis (SBS) chemistry, which delivers the highest yield of error-free reads and the highest percentage of base calls above Q30 in the industry. For larger projects, the high-output mode uses 8 lane flow cells, which produce 1.5 billion reads at speeds of 1 and 6 days for 1 x 50-cycle runs and 2 x 125-cycle, respectively. An optional rapid-run mode integrates onboard clustering, a unique two-lane flow cell, and faster cycle times, generating 300 million reads per flow cell with a turnaround time of 60 hours for a 2 X 250 paired-end run. Only 12 hours of runtime is required for 50-cycle short reads, such as differential gene expression analysis or ChIP-seq. This new feature will benefit researchers who typically desire single-lane sequencing since there is no wait time to fill a high-output flow cell. The Hiseq 2500’s speed, length of reads, and throughput make it a good choice for a wide range of sequencing applications.

IMPORTANT: This equipment was funded by NIH equipment grant S10OD016262. Please reference this award in any publications that utilize this equipment.

Illumina Miseq

The baby brother to the popular Hiseq. It also uses Illumina sequencing by synthesis and can complete a 500 cycle paired-end run in a couple of days on its one-lane flowcell. Depending on the type of library, it can produce anywhere from 7M to 16M reads on a full Flowcell kit. If you are interested in a smaller amount of reads, Illumina also offers Nano and Micro version kits. The most popular applications for the Miseq are enriched/targeted samples, bisulfite treated, and amplicons.

Agilent 2200 Tapestation

The Tapestation is an automated electrophoretic-based nucleic acid / protein analyzer that is indispensible for QC steps in library prep for NGS. In comparison to Bioanalyzer, the Tapestation provides savings on consumables, faster and more accurate data, and is capable of high throughput processing of 96-well plates. The automated loading reduces user error and the potential to waste precious samples. The pre-cast gel strips provide scalable throughput at a constant cost per sample, therefore it makes an excellent solution for high throughput and also eliminates unnecessary costs associated with low sample numbers.

Agilent Bioanalyzer

Although our preferred method for DNA/RNA analysis is the Tapestation because of its cost reduction and ease of use, the Bioanalyzer is still a workhorse in our RNA library preparation pipeline. It only uses 1ul, which saves on precious samples, and it is much more sensitive for low concentrations.

Covaris model E220

Currently, Covaris technology is the gold standard for fragmenting DNA in order to get a consistently tight size range. Although we use the Covaris E220 primarily for Chromatin shearing and fragmenting DNA in our library preparation pipeline, it has many other uses. The instrument achieves a high level of consistency and efficiency through its use of isothermal processing and the proprietary Covaris Adaptive Focused Acoustic technology. It can process 1 to 96 samples in a single batch, which is much more efficient than using our S2 model for large arrays of samples.

Covaris model E220

Currently, Covaris technology is the gold standard for fragmenting DNA in order to get a consistently tight size range. Although we use the Covaris E220 primarily for Chromatin shearing and fragmenting DNA in our library preparation pipeline, it has many other uses. The instrument achieves a high level of consistency and efficiency through its use of isothermal processing and the proprietary Covaris Adaptive Focused Acoustic technology. It can process 1 to 96 samples in a single batch, which is much more efficient than using our S2 model for large arrays of samples.

Beckman Coulter Biomek FXP

The La Jolla Institute is currently engaged in a partnership with Beckman Coulter and Illumina for the purposes of developing and validating Illumina library preparation methods on the Biomek FXP automation platform. Currently, we have the Truseq mRNA Stranded, Truseq Nano DNA and Nextera based protocols installed on the system. Implementing automation for library preparation should reduce inconsistency and risks associated with human errors. The ability to generate libraries, either from a single sample to as many as 96 at a time, will benefit both small-scale and particularly large-scale experiments. We wish to keep a long-standing intimate relationship with both Illumina and Beckman Coulter to continue improving our and other’s NGS needs.

Rates

  2018 Internal Rates 2018 External Rates
RNA extraction
Up to 24 samples (Qiagen miRNAesay micro kit)/sample $21.00 $39.69
24-48 samples (Zymo 96 magnetic kit, automated)/sample $18.00 $34.02
49-96 samples (Zymo 96 magnetic kit, automated)/sample $10.00 $18.90
External NP Usage fee per Sample $5.00 $9.45
Agilent Tape Station (per 16 samples)
D1K / HS DNA Screen Tape - DIY $45.00 n/a
D1K / HS DNA Screen Tape - Full Service $105.00 $198.45
R6K / HS R6K RNA Screen Tape - DIY $45.00 n/a
R6K / HS R6K RNA Screen Tape - Full Service $105.00 $198.45
External NP Usage Fee $40.00 $75.60
Agilent Bioanalyzer (per 11 or 12 samples)
DNA 1000 Chip (12) - DIY $32.00 n/a
DNA 1000 Chip (12) - Full Service $92.00 $173.88
HiSensitivity DNA Chip (11) - DIY $50.00 n/a
HiSensitivity DNA Chip (11) - Full Service $110.00 $207.90
RNA 6000 Nano Chip (12) or Pico Chip (11) - DIY $32.00 n/a
RNA 6000 Nano Chip (12) or Pico Chip (11) - Full Service $92.00 $173.88
External NP Usage Fee $40.00 $75.60
Illumina Library Preparation
TruSeq Stranded mRNA - DIY $70.00 n/a
TruSeq Stranded mRNA - Full Service (1 to 12 samples) $160.00 $302.40
TruSeq Stranded mRNA - Full Service additional per sample (13-96) $85.00 $160.65
TruSeq Nano DNA prep - DIY $48.00 n/a
TruSeq Nano DNA prep - Full Service (1 to 12 samples) $108.00 $204.12
TruSeq Nano DNA prep - Full Service additional per sample (13-96) $63.00 $119.07
Nextera DNA library preps - DIY $44.00 n/a
Nextera DNA library preps - Full service $74.00 $139.86
Nextera - Full Service additional per sample (13-96) $59.00 $111.51
External NP Usage Fee $40.00 $75.60
Library Pooling and QC
Library pooling and pre-run QC (up to 16 samples) $200.00 $378.00
Library pooling and pre-run QC Additional per sample beyond 16 $5.00 $9.45
External NP Fee $100.00 $189.00
Illumina Sequencing (Rapid Run Flowcell, 2 lanes )
Rapid Run Single End 50bp reads (SE50) $1,645.60 $3,110.18
Rapid Run Single End 100bp reads (SE100) $2,181.20 $4,122.47
Rapid Run Paired End End 50bp reads (PE100) $2,775.20 $5,245.13
Rapid Run Paired End 100bp reads (PE200) $3,654.00 $6,906.06
Rapid Run additional 50-cycles $600.00 $1,134.00
Rapid Run External usage fee SE per run $500.00 $945.00
Rapid Run External usage fee PE per run $750.00 $1,417.50
Rapid Run Single End HT genomics users with lab supplied reagents $200.00 n/a
Rapid Run Paired-End HT genomics users with lab supplied reagents $300.00 n/a
Illumina Sequencing (V4 High Output Flowcell, 8 lanes )
High Output Run - Single End 50bp reads (SE50) $7,132.08 $13,479.63
High Output Run - Single End 100bp reads (SE100) $9,689.44 $18,313.04
High Output Run - Paired End 50bp reads (PE100) $12,071.44 $22,815.02
High Output Run - Paired End 125bp reads (PE250) $16,857.52 $31,860.71
High Output External usage fee SE per flowcell $800.00 $1,512.00
High Output External usage fee PE per flowcell $1,200.00 $2,268.00
High Output Run - Single End HT genomics users with lab supplied reagents $500.00 n/a
High Output Run - Paired End HT genomics users with lab supplied reagents $750.00 n/a
cBOT Rapid Run Duo Lane splitting $600.00 $1,134.00
Illumina Sequencing (Miseq )
MiSeq Full Service (does not include reagents) $200.00 $378.00
External NP Usage Fee $200.00 $378.00
DNA Shearing (Covaris E-Series)
Covaris Access-per sample (includes tube) - DIY $5.00 $9.45
Covaris usage fee (/hour) (w/o cooling) $60.00 $113.40
Technical Assistance per hour $60.00 $113.40
Other
Technical Training (Sequencing specialist) / hour $60.00 $113.40
Scientific consultation /hour (first meeting free of charge) $90.00 $170.10