Here are our favorite resources about next generation sequencing (NGS) as well as information about the sequencing platforms we house. If you’re new to NGS, please check out this beginners guide.
We measure DNA concentrations with Qubit and band sizes for possibly degraded DNA with the Agilent Bioanalyzer. Here is quick refresher on genomic DNA quantity and quality.
*Please note: NanoDrop Spectrophotometers can overestimate concentrations. Take this into consideration if submitting samples that have been quantified this way.
For Illumina sequencing, we recommend the following kits for genomic DNA extractions:
For long read sequencing, we recommend the following kits for high molecular weight genomic DNA extractions:
We measure RNA concentrations with Qubit and band sizes with the Agilent Bioanalyzer. Here is a quick refresher on RNA quality. We strongly recommend DNase treatment on RNA samples.
*Please note: NanoDrop Spectrophotometers can overestimate concentrations. Take this into consideration if submitting samples that have been quantified this way.
We recommend the following kits for RNA extractions:
Short Read Sequencing
Illumina’s Next Generation Sequencing technology uses sequencing by synthesis (SBS) chemistry and allows for both single-read and paired-end sequencing options up to 300 bp in length.
We offer short read sequencing services on our Illumina NovaSeq X Plus, MiSeq, and NextSeq 1000 sequencers. As all Illumina sequencers use similar chemistry, same library preparations can be used across these instruments, allowing us to tailor to researchers’ sequencing needs according to read length, throughput, and turnaround time. Illumina provides sequencing coverage recommendations to help determine how many reads are needed for an experiment.
Multiple samples can be indexed by a unique nucleotide sequence and pooled into an individual run to save costs. Samples may be indexed using Illumina’s TruSeq indices, Nextera indices, or custom indexing adaptors. The SGT can demultiplex the reads and provide fastq files for each individual sample if Illumina indices are used.
*Please note: If submitting customer prepared libraries, each library must contain a unique index barcode to allow for pooling and accurate demultiplexing. Indexes cannot be reused for libraries in the same pool.
The NovaSeq X Plus is a the latest robust and flexible, production-scale sequencer from Illumina that utilizes patterned flow cells and novel 2-color channel XLEAP chemistry. XLEAP chemistry reduces reagent costs and decreases data processing times while still producing the same high-quality data. The NovaSeq X also allows the flexibility to sequence by flow cell or lane.
SEQUENCING OUTPUT PER FLOW CELL
Product | # of Cycles | Total Reads per Flow Cell | Total Reads per Lane | Output (Gb) | Duke Price | Non-Profit External Price |
---|---|---|---|---|---|---|
1.5B flow cell |
100 Cycles |
1.5 B | n/a | 150 | $2,589 | $2,999 |
1.5B flow cell | 200 Cycles (100x100) |
1.5B | n/a | 300 | $3,139 | $3,576 |
1.5B flow cell | 300 Cycles (150x150) |
1.5 B | n/a | 450 | $3,359 | $3,807 |
1.5 B lane | Any Config | n/a | 700 M | variable | $1,929 | $2,306 |
10B flow cell | 100 Cycles (50x50) |
10 B | n/a | 1,000 | $10,798 | $13,012 |
10B flow cell | 200 Cycles (100x100) |
10 B | n/a | 2,000 | $12,888 | $15,206 |
10B flow cell | 300 Cycles (150x150) |
10 B | n/a | 3,000 | $13,548 | $15,899 |
10B lane | Any Config | n/a | 1 B | variabl | $1,818 | $2,188 |
25B flow cell | 300 Cycles (150x150) |
25 B | n/a | 8,000 | $22,832 | $26,904 |
25B lane | Any Config | n/a | 3 B | variable | $2,872 | $3,504 |
The MiSeq is a benchtop sequencer from Illumina that allows for more focused applications, such as targeted gene sequencing, metagenomics, small genome sequence, targeted gene expression, and amplicon sequencing. The ability to run long paired-end reads (2 x 250bp or 2 x 300bp) makes this instrument ideal for amplicon sequencing. The MiSeq is capable of generating ~12 million reads passing filter for v2 kits (~24 million for paired-end sequencing) and ~22 million reads passing filter for v3 kits (~44 million for paired-end sequencing).
Flow Cell | # of Cycles | Total Reads per Flow Cell | Total Reads per Lane | Output (Gb) | Duke Price | Non-Profit External Price |
---|---|---|---|---|---|---|
MiSeq NANO V2 |
300 Cycles |
1 M | n/a | 0.3 | $808 | $1,088 |
MiSeq NANO V2 | 500 Cycles (250x250) |
1 M | n/a | 0.5 | $886 | $1,169 |
MiSeq MICRO V2 | 300 Cycles (150x150) |
4 M | n/a | 1.2 | $1,004 | $1,293 |
MiSeq V2 | 50 Cycles (50 SR) |
12 M | n/a | 0.75 | $1,500 | $1,814 |
MiSeq V2 | 300 Cycles (150x150) |
12 M | n/a | 4.5 | $1,802 | $2,131 |
MiSeq V2 | 500 Cycles (250x250) |
12 M | n/a | 6.0 | $1,973 | $2,311 |
MiSeq V3 | 150 Cycles (75x75) |
22 M | n/a | 3.3 | $1,619 | $1,939 |
MiSeq V3 | 600 Cycles (300x300) |
22 M | n/a | 13.2 | $2,439 | $$2,800 |
The NextSeq 1000 is a new benchtop sequencer from Illumina that utilizes a patterned flow cell, 2-color channel chemistry, and an integrated sequencing cartridge. The patterned flow cell features a novel, high resolution optical system and higher cluster densities to allow for increased data output with greater resolution. Two flow cell types are offered to help cater to coverage needs: P1 and P2.
Flow Cell | # of Cycles | Total Reads per Flow Cell | Total Reads per Lane | Output (Gb) | Duke Price | Non-Profit External Price |
---|---|---|---|---|---|---|
NextSeq 1000 XLEAP P1 | 100 Cycles (50x50) |
100 M | n/a | 10 | $1,227 | $1,533 |
NextSeq 1000 XLEAP P1 | 300 Cycles (150x150) |
100 M | n/a | 30 | $1,535 | $1,862 |
NextSeq 1000 XLEAP P1 | 600 Cycles (300x300) |
100 M | n/a | 60 | $1,956 | $2,457 |
NextSeq 1000 XLEAP P2 | 100 Cycles (50x50) |
400 M | n/a | 40 | $1,637 | $1,964 |
NextSeq 1000 XLEAP P2 | 200 Cycles (100x100) |
400 M | n/a | 80 | $@,694 | $3,073 |
NextSeq 1000 XLEAP P2 | 300 Cycles (150x150) |
400 M | n/a | 120 | $3,430 | $3,846 |
NextSeq 1000 XLEAP P2 | 600 Cycles | 400 M | n/a | 240 | $3,695 | $4,124 |
Flow Cell | # of Cycles | Total Reads per Flow Cell | Total Reads per Lane | Total GBP | Duke Price | Outside of Duke Price |
---|---|---|---|---|---|---|
MiSeq NANO V2 | 300 Cycles (150x150) | 1 M | n/a | 0.3 | $808 | $1088 |
MiSeq NANO V2 | 500 Cycles (250x250) | 1 M | n/a | 0.5 | $886 | $1,169 |
MiSeq MICRO V2 | 300 Cycles (150x150) | 4 M | n/a | 1.2 | $1004 | $1,293 |
MiSeq V2 | 50 Cycles (50 SR) |
12 M | n/a | .75 - .85 | $1,500 | $1,814 |
MiSeq V2 | 300 Cycles (150x150) | 12 M | n/a | 4.5 | $1,802 | $2,131 |
MiSeq V2 | 500 Cycles (250x250) | 12 M | n/a | 6.0 | $1,973 | $2,311 |
MiSeq V3 | 150 Cycles (75x75) | 22 M | n/a | 3.3 | $1,619 | $1,939 |
MiSeq V3 | 600 Cycles (300x300) | 22 M | n/a | 13.2 - 15 | $2,439 | $2,800 |
NextSeq 1000 P1 | 100 Cycles (50x50) | 100 M | n/a | 10 | $1,227 | $1,533 |
NextSeq 1000 P1 | 300 Cycles (150x150) | 100 M | n/a | 30 | $1,535 | $1,862 |
NextSeq 1000 P1 | 600 Cycles (300x300) | 100 Million | n/a | 60 | $1,956 | $2,457 |
NextSeq 1000 P2 | 100 Cycles (50x50) | 400 M | n/a | 40 | $1,637 | $1,964 |
NextSeq 1000 P2 | 200 Cycles (100x100) | 400 M | n/a | 80 | $2,694 | $3,073 |
NextSeq 1000 P2 | 300 Cycles (150x150) | 400 M | n/a | 120 | $3,430 | $3,846 |
NextSeq 1000 P2 | 600 Cycles (300x300) | 400 M | n/a | 240 | $3,695 | $4,124 |
NovaSeq X 1.5B | 100 Cycles (50x50) | 1.5 B | n/a | 150 | $2,589 | $2,999 |
NovaSeq X 1.5B | 200 Cycles (100x100) | 1.5 B | n/a | 300 | $3,139 | $3,576 |
NovaSeq X 1.5B | 300 Cycles (150x150) | 1.5 B | n/a | 450 | $3,359 | $3,807 |
NovaSeq X 1.5B | Any Config | n/a | 750 M | variable | $1,929 | $2,306 |
NovaSeq X 10B | 100 Cycles (50x50) | 10 B | n/a | 1,000 | $10,798 | $13,012 |
NovaSeq X 10B | 200 Cycles (100x100) | 10 B | n/a | 2,000 | $12,888 | $15,206 |
NovaSeq X 10B | 300 Cycles (150x150) | 10 B | n/a | 3,000 | $13,548 | $15,899 |
NovaSeq X 10B | Any Config | n/a | 1 B | variable | $1,818 | $2,188 |
NovaSeq X 25B | 300 Cycles (150x150) | 25 B | n/a | 8,000 | $22,832 | $26,904 |
NovaSeq X 25B | Any Config | n/a | 3 Billion | variable | $2,872 | $3,504 |
Long Read Sequencing
Long read sequencing, also known as third generation sequencing, involves reading sequences that are thousands of bases long. The longer read length generates a reasonable amount of overlap between sequences for better sequence assembly. We offer long read sequencing on the PacBio Revio sequencer.