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Sequencing Platform

 

 

 

 

 

The sequencing platform at the LCSB offers next-generation sequencing services. The platform has a NextSeq500 and a MiSeq sequencer from Illumina, and a MinION nanopore sequencer from Oxford Nanopore Technologies, which allow for high flexibility when it comes to different sequencing needs.
For first-time users as well as for researchers with new projects, we encourage to meet in person and discuss the project objectives and goals. It helps us understand your needs, which are almost unique for every project. Following our discussion, we offer our best advice and recommendations related to best practices to obtain the relevant and good quality sequencing data. Some of the points that we suggest considering for your sequencing projects are:

  • How many samples (number) are necessary to obtain a robust result?
  • How are these samples collected?
  • Which DNA/RNA isolation technique is optimal?
  • How is better RNA integrity achieved?
  • How many biological replicates are necessary?
  • How much sequencing data coverage (depth) is required?

 

Equipment

MiSeq™
(Illumina)

 

 

MiSeq™ is a tabletop sequencer from Illumina®, Inc. and is well suited for sequencing projects such as metagenomics, small genome sequencing, targeted resequencing and targeted gene expression profiling.

The applications can be tuned to individual requirements as the MiSeq™ system along with its different reagent kits can offer up to 25 million reads per run and up to 15 GB of sequencing data, and up to 2 x 300 bp read length as well.

NextSeq500™
(Illumina)

 

 

NextSeq500™ is a tabletop sequencer from Illumina®, Inc. and is well suited for sequencing projects requiring whole-genome sequencing, exome sequencing, whole-transcriptome sequencing, or targeted gene sequencing.

With a higher throughput than MiSeq™ sequencer, NextSeq500™ sequencer along with its different reagent kits can offer up to 400 million reads per run and up to 120 GB of sequencing data. The maximum read can be 2 x 150 bp.

MinION®
(Oxford Nanopore)

 

 

MinION® is a sequencer from Oxford Nanopore Technologies and offers the possibility to sequence DNA or RNA fragments of versatile lengths, even up to several hundred kilobases long fragments.

MinION® sequencer is well suited for sequencing projects interested in de novo sequencing, targeted sequencing, metagenomics, transcriptome and epigenetics. It can generate up to 20 GB of data per flow cell (48 hrs of sequencing run).

Services

Sample Quality Control: We offer sample quality control for both quantity and quality as complimentary service. Please be advised that if the sample quality is suspected below normal standards, the procedure will be stopped.

Sequencing Run: We offer the users with the possibility to provide us with their samples (DNA or RNA) or prepared sequencing libraries.

We have strong expertise in sequencing samples for the following applications:

Short read sequencing

  • Transcriptomics:
    • Bulk RNA sequencing
    • Single cell sequencing
    • Meta-transcriptomics sequencing
  • Genomics:
    • Whole genome sequencing (small genomes)­
    • Chromatin immunoprecipitation sequencing (ChIP-Seq)
    • Methylated DNA immunoprecipitation sequencing (MeDIP-Seq)
    • Reduced representation bisulfite sequencing (RRBS)
    • Assay for Transposase-Accessible Chromatin sequencing (ATAC-Seq)
    • Exome-sequencing
    • Meta-genomics
  • Targeted Resequencing:
    • RNA
    • DNA
    • DNA methylation
  • Small RNA sequencing

Long read sequencing:

  • mtDNA sequencing
  • genomic DNA sequencing
  • full length 16S/18S rRNA sequencing
  • full length cDNA sequencing

In case you have a project with an application not listed above, please contact us regarding the feasibility.

Data Analysis

Upon request, the LCSB sequencing platform offers the following primary data analysis:

  • The fastq files after demultiplexing (sequence read data).
  • Quality control report for using the fastQC software.

For advanced secondary analysis, we recommend contacting the Bioinformatics Core and the Eco-Systems Biology group (for metagenomics/meta-transcriptomics analysis using IMP). These services are separate to that of LCSB sequencing platform and therefore, has to be inquired independently by the user for any extra charges and/or authorship(s).

Sequencing

Turn Around Time: Following the sample submission to the LCSB sequencing platform and fulfilment of sample and administrative requirements, the normal turn-around time is between 2 to 3 weeks. However, we may have either a large set of samples in process queue or unforeseen situations like unscheduled instrument maintenance, etc., which may delay our planned sample processing. In these cases, we notify via e-mail.

Data Transfer: Once the sequencing run is completed, the sequencing data is available to University Luxembourg researchers with access to internal computational resources via folder sharing (group folder on Isilon).

For other users, we offer data transfer via (i) secured FTP transfer, or (ii) secured folder accessibility or (iii) user provided solution - like user Onedrive, ownCloud etc.

Data Storage: The LCSB sequencing platform does not offer any long-term data storage and to continuously make space for new data, the platform typically deletes the sequencing data in 4 weeks following the data delivery. It is therefore the sole responsibility of the user to keep the primary (unprocessed) sequencing data for long term storage. We also highly recommend to archive the sequencing data at multiple independent sites/folders.

 

How to use

We offer a broad range of services and to get a personalised quote, please contact us with the following details:

  • Number of samples.
  • Sequencing depth (expected GB of data per sample) needed.
  • Expected time-frame of availability of samples.
  • User and PI affiliation (institution).

Please let us know if the proposed project is exploratory in nature or a pilot study, as we understand the associated risks of such projects and offer discounted prices for limited number of samples.

PS: Please note that the final invoice for our services are only available electronically (via e-mail) and would not be sent via normal post unless specifically asked for.

Forms

Project Information Form

Illumina Sequencing Forms

Nanopore Sequencing Forms

Service Terms and Contract

Requirements [A + B]

It is necessary to fulfil both the sample (A) and administrative (B) requirements before the LCSB sequencing platform can start processing the samples. It is therefore advisable to follow-up on both the requirements simultaneously.

Please note that during our quality control, if the sample quality is suspected below normal standards, the procedure will be stopped.

Sample Requirements [A = A.1 + A.2 + A.3 + A.4]

Sample requirement for Illumina – DNA

Sample Quantity (A.1)

  • Genomic DNA – 1 µg (recommended)
  • ChIP DNA – 10 ng (minimum)
  • Sample volume – 10 µl (minimum)

Sample Quality (A.2)

  • OD260/280 ≃ 1.8
  • OD260/230 = (1.8 – 2.0)
  • If possible, please provide Bioanalyzer results

Sample Preparation (A.3)

  • Use same extraction protocol for all samples
  • DNA should be RNase treated
  • DNA should be resuspended in
    • nuclease free water, or
    • lowTE (10 mM Tris, 0.1 mM EDTA)

Sample Labelling (A.4)

  • Use only 1.5 ml eppendorf tubes
  • Use black or blue permanent markers for labelling
  • Do not use special characters in sample names
  • Transport DNA at 4℃ or frozen

Sample requirement for Illumina – RNA

Sample Quantity (A.1)

  • Total RNA – 2 µg (minimum)
  • Sample volume – 10 µl (minimum)

Sample Quality (A.2)

  • OD260/280 ≃ 2.0
  • OD260/230 = (1.8 – 2.0)
  • Agilent Bioanalyzer RIN values > 7.0
  • Ratio of high/low MW rRNA ~2

Sample Preparation (A.3)

  • Use same extraction protocol for all samples
  • RNA should be DNase treated
  • RNA should be resuspended in RNase free water

Sample Labelling (A.4)

  • Use only 1.5 ml eppendorf tubes
  • Use black or blue permanent markers for labelling
  • Do not use special characters in sample names
  • Transport RNA only on dry ice

Sample requirement for Illumina – Library

Library Quantity (A.1)

  • Library concentration - 10nM (minimum)
  • Sample volume – 10 µl (minimum)

Library Quality (A.2)

  • Bioanalyzer high sensitivity profile or similar

Library Preparation (A.3)

  • Final Library should be in lowTE (10mM Tris, 0.1 mM EDTA)

Library Labelling (A.4)

  • Use only 1.5 ml Eppendorf tubes
  • Use black or blue permanent markers for labelling
  • Do not use special characters in sample names
  • Transport Library(s) only on dry ice

Sample requirement for Nanopore – DNA

Sample Quantity (A.1)

  • Genomic DNA – 5->15 µg (depending upon the DNA size)
  • PCR based method – 10ng
  • Sample volume – 20 µl (minimum)

Sample Quality (A.2)

  • OD260/280 ≃ 1.8
  • OD260/230 = (1.8 – 2.0)
  • Gel or Bioanalyzer profile or similar

Sample Preparation (A.3)

  • Use same extraction protocol for all samples
  • DNA should be RNase treated
  • DNA should be resuspended in
    • nuclease free water, or
    • lowTE (10 mM Tris, 0.1 mM EDTA)

Sample Labelling (A.4)

  • Use only 1.5 ml eppendorf tubes
  • Use black or blue permanent markers for labelling
  • Do not use special characters in sample names
  • Transport DNA at 4℃ or frozen

Administrative Requirements [B = B.1 + B.2 + B.3]

Researchers using Luxembourg University OTP codes

  • Completed Sample Information Form (B.1): The correct sample information form(s) must be completed and returned via e-mail.
  • Confirmation of Funds Availability (B.2): It is not a mandatory step but highly advisable that the researcher confirms availability of necessary funds in their respective OTP codes to avoid administrative issues later, for example, the expiry/maturity dates of the OTP code(s), etc.
  • Agreement to the 'General Terms and Conditions' (B.3): Upon sample submission to the LCSB sequencing platform, it is assumed that the researcher has read and agreed to the 'General Terms and Conditions' for the services provided by the sequencing platform.

Researchers using other budgets

  • Completed Sample Information Form (B.1): The correct sample information form(s) must be completed and returned via e-mail.
  • A 'Purchase Order' or signed agreement of 'Service Contract' (B.2): It is compulsory to provide either a 'Purchase Order' or a signed 'Service Contract' via e-mail. To raise a 'Purchase Order' of the correct amount, please contact us for a quotation.
  • Agreement to the 'General Terms and Conditions' (B.3): Upon sample submission to the LCSB sequencing platform, it is assumed that the researcher has read and agreed to the 'General Terms and Conditions' for the servives provided by the sequencing platform.

 

Contact information

Reach Us
E-mail: LCSB-sequencing [at] uni.lu
Phone: (+352) 466 644 6189

Shipping Address
Sequencing Platform
BioTech I - Luxembourg Centre for Systems Biomedicine
Campus Belval, Université du Luxembourg
7, Avenue des Hauts Fourneaux - L-4362 Esch-sur-Alzette
Location on Google Maps ⇗