Next Generation Services
Next-generation sequencing (NGS), also called massively parallel or deep sequencing, stands for a DNA/RNA sequencing technology that is much quicker and cheaper than the previously used Sanger sequencing and has revolutionized genomic research. A good summary for users new to NGS technologies can be found in the document "An Introduction to next generation technology". Please see below for an overview of our services. You can book all our services via iLab.
Next-generation sequencing (NGS), also called massively parallel or deep sequencing, stands for a DNA/RNA sequencing technology that is much quicker and cheaper than the previously used Sanger sequencing and has revolutionized genomic research. A good summary for users new to NGS technologies can be found in the document "An Introduction to next generation technology". Please see below for an overview of our services. You can book all our services via iLab.
Service Overview
Our NGS platform provides sequencing services for both DNA and RNA analysis as follows:
- Automated DNA/RNA Isolation
- Manuell and automated library preparation for WGS, Exome-Seq, and RNA-Seq (mRNA and total RNA)
- NGS Sequencing full service
- Sequencing only service (users provide Illumina kit)
- Method development
- Consultation on experimental design and troubleshooting
- Support for the submission of grants and publications
NGS services
For high throughput, standardized library preparation we use three Agilent Bravo NGS workstations (pre and post PCR) to process up to 96 samples per run.
Our library prep portfolio:
- whole genome library preparation for DNA (Illumina PCR-free tagmentation kit)
- whole exome library preparation for DNA (Twist Exome/Panels)
- mRNA and totalRNA library preparation (Illumina Stranded mRNA Ligation and Illumina Stranded Total RNA Ligation with Ribo-Zero Plus)
Crucially important for successful and high quality library prep is the exact measurement of the concentration and the quality of the submitted DNA/RNA. Input DNA and RNA quantities specified in table 1 apply only if samples were quantified by a fluorometric method (e.g. Qubit, PicoGreen, RiboGreen). If a spectrophotometer (e.g. Nanodrop) was used, we suggest submitting twice the requested amount of sample since this type of measurement is often unreliable. In any case, sample amounts higher than the minimum requirements will improve the library complexity and thus the quality of the data.
Prepared libraries are ready to be sequenced on the NextSeq 1000 or NovaSeq X Plus depending on the desired output. If you have a large project that would require a different library preparation method, get in contact with us to discuss the automation possibilities.
Table 1: Minimum amount and quality of nuclic acids for various library prep protocols
Library Type | Quality | Quantity** | Concentration | Buffer |
Whole Genome Sequencing | double-stranded DNA, non-degraded, A260/280 ratio > 1.8 | 500 ng | 20 ng/µl | EB*, H2O |
DNA PCR-Free Prep, Tagmentation library | checked with agarose gel, RNA-free | in 25 µl | 20 ng/µl | EB*, H2O |
Twist Exome/Panels | double-stranded DNA, non-degraded, A260/280 ratio > 1.8, RNA-free | 65 ng in 13 µl | 5 ng/µl | EB*, H2O |
Stranded mRNA, Ligation | mRNA, RIN > 7, DNA-free | 300 ng - 1 µg in 25 µl | 12-40 ng/µl | non-DEPC treated H2O |
Stranded Total RNA, Ligation | Total RNA, RIN > 7, DNA-free | 10 - 200 ng in 11 µl | 0.9-19 ng/µl | non-DEPC treated H2O |
*EB: Elution Buffer, **Measured with a fluorescence-based method like Qubit
The Illumina NextSeq 1000 sequencing system can run two different flow cell types (P1 and P2) and various read length combinations to enable effective throughput as needed (100 Mio reads in P1 and 400 Mio reads in P2). This benchtop sequencer creates the perfect output bridge between the low throughput Miseq sequencer (up to 25Mio reads) and the high throughput NovaSeq 6000 (starting with 800 Mio reads). Flow cell features are summarized in tables 1 and 2 below.
Table 1: Sequencing Output Per Flow CellNextSeq 1000
Flowcell type | P1 | P2 |
2 x 50 | N/A | 40 Gb / 400 Mio reads |
2 x 100 | N/A | 80 Gb / 400 Mio reads |
2 x 150 | 30 Gb / 100 Mio reads | 120 Gb / 400 Mio reads |
Table 2: Estimated Sample Throughput for Key Applications
Flow Cell Type | P1 | P2 |
Small Whole-Genome Sequencing (300 cycles) | ~7 | ~30 |
Whole-Exome Sequencing (200 cycles) | ~4 | ~16 |
Single-Cell RNA-Seq (100 cycles) |
| ~2-10 |
Source: https://emea.illumina.com/systems/sequencing-platforms/nextseq-1000-2000/specifications.html
We offer the services below for sequencing on the Nextseq 1000, routinely the core will order all reagents and invoice for the complete service:
- Sequencing plus library-Prep from isolated DNA/RNA
- Sequencing of customer-made libraries
- Sequencing only service (Customer delivers Illumina Sequencing Kits and ready made library)
The Illumina Novaseq X Plus sequencing system is the state-of-the-art sequencer for genomic studies.
It can run three types of Flowcells (1.5B, 10B and 25B). These flowcells and various read length combinations enable effective throughput as needed. Flow cell features are summarized in tables 1 and 2 below.
The core will determine the best solution with each customer.
Table 1: Flowcell-Options for the Illumina NovaSeq X Plus
Flow Cell Type | 1.5B | 10B | 25B |
Available sizes (In cycles) | 100 200 300 | 100 200 300 | 100 200 300 |
Single or Paired-reads | 1.5 Billions | 10 Billions | 25 Billions |
Number of lanes | 2 | 8 | 8 |
Please note: The NovaSeq X Plus Kits contains reagents for 38 additional cycles to support the use of Unique Dual Indexes (UDIs) and Unique Molecular Identifiers (UMIs).
Table 2: Estimated sample throughput for frequently used applications
Flow Cell Type | 1.5B | 10B | 25B |
Human Genomes per Flow Cell | ~4 | ~24 | ~64 |
Exomes per Flow Cell | ~41 | ~250 | ~750 |
Transcriptomes per Flow Cell | ~30 | ~200 | ~520 |
*All sample throughputs are estimates per single flow cell run. Human genomes assumes > 120 Gb of data per sample to achieve 30× genome coverage. Exomes assumes ~8 Gb per sample to achieve 100x coverage. Transcriptomes assumes ≥ 50M reads. Throughput may vary based on library preparation kit used.
Source: https://sapac.illumina.com/systems/sequencing-platforms/novaseq-x-plus/specifications.html
We offer the options below for sequencing on the NovaSeq, routinely the core will order all reagents and invoice for the complete service:
Sequencing plus library-Prep from isolated DNA/RNA
Sequencing of customer-made libraries
Sequencing only service (Customer delivers Illumina Sequencing Kits and ready made library)
- Illumina NovaSeq X Plus
- Illumina NextSeq 1000
- 10x Genomics Chromium
- Agilent Bravo robotic workstations (library prep)
- TECAN Fluent robotic workstation
- Perkin Elmer LabChip GX Touch HT
- QC-Instruments (Qubit, Nanodrop,Covaris200)
