Genomics Core Library Preparation Submission

Library Prep Reagent Kit Submit Submission form workflow Quantity range (ng) Volume required per prep (µl)* Sample Batch Size** Notes
Illumina Stranded mRNA Total RNA Illumina stranded mRNA 25 - 1000 25 Any number This kit supports various input mass; normalise where possible.
NEBNext Single cell / low input RNA Library Prep Kit for Illumina Total RNA / cells NEB low input RNA 2pg - 200ng, or cell(s) 8 24 Please discuss your input requirements prior to submission.
Illumina DNA PCR-Free Library Prep, Tagmentation Genomic DNA Illumina DNA PCR Free, Tag 25 - 2000 30 24 This kit provides a "low-input" protocol for small input mass. It will also self-normalise samples above 300 ng.
Illumina DNA Prep with Enrichment Genomic DNA Illumina DNA Prep with Enrichment 100 - 500 30 94 Also suitable for FFPE extracted material using a modified protocol.
ThruPLEX Tag-Seq HV ChIP'ed DNA Thruplex Tag Seq 5 - 200 30 24 This kit uses unique molecular identifiers.
Illumina TruSeq RNA HT (Total) Total RNA Illumina Total RNA 1 - 1000 11 16 Also suitable for FFPE extracted material using a modified protocol.

* Please provide an additional 15% volume per well.

** We run a positive and negative control with every library prep.

Library Prep Submission Process

  1. 96 well barcoded plates and seals are available from the Genomics Core (Room 026).
  2. Prepare your samples in columns following the concentration and volumes in the table above.
  3. If applicable, plate your samples according to the randomisation layout provided by your bioinformatic contact (discussed at your experimental design meeting).
    • Best practice is to at least randomise your plate layout so sample groups are not co-located by row or column.
  4. The bioinformatics contact on your project can confirm the quantity of sequence data required, read length, and recommended sequencer. This translates to the number of flowcell lanes required and run parameters.
  5. Complete the submission form:
    • The Plate ID field is the barcode from the submission plate.
    • Set Flowcell Type only if submitting for NovaSeq; leave the field empty for all other sequencers.
    • Complete Billing Information. This is essential for correct invoicing: please follow the popup instructions in the field. Only enter a PO Number if required by your department or organisation. Leave PO Number blank if it does not apply (resist the temptation to put "N/A" or "none": it will complicate your department's invoice).
    • Ensure the sample names match those provided to the Bioinformatics Core (if applicable) and well position (Row and Column) match the plate layout.
    • The RIN field is only relevant for RNA submissions.
    • Leave unused sample rows empty.
    • You may submit more than one uniquely barcoded 96 well plate by copying the first sheet of the submission workbook as an additional sheet in the workbook. You can make as many sheets as you need, filling in each sheet with exactly one additional uniquely barcoded 96 well plate.
  6. Run your completed submission form through this submission application. Be sure to click the "Confirm Submission" button to finish the submission process.
  7. Hand your plate to a member of Genomics. RNA plates are stored at -80°C until processing.

Library Prep Submission Recommendations

  1. When eluting or diluting your sample, avoid using buffers containing high concentrations of EDTA and use DNAse as recommended in the kit manufacturer's instructions for RNA extraction.
  2. Quantify your nucleic acid using a fluorescent based method.
  3. When submitting DNA, we recommend performing QC to include size information using the Agilent Tape Station or equivalent.
  4. When submitting RNA, check the quality of each sample on the Bioanalyzer or Tapestation. If the RIN is <8, you can contact the Genomics Core for advice. A low quality score won't necessarily stop it being processed but can certainly lead to poor sequencing results, it is better to have this information if possible.
  5. For any ChIP-seq experiment, you will have performed the chromatin immunoprecipitation and just submit the ChIP'ed DNA.
  6. When submitting ChIP'ed DNA, check the fragmentation profile of your input(s) material. Please discuss with the Genomics Core if the fragments appear larger than 700bp.
  7. We do not specify a minimum quantity for a ChIP-seq experiment; please submit what you have. Always include at least one input control.
  8. Please upload QC data to the relevant project in our LIMS.

Hints and Tips

  1. This is the submission form for library prep specifically. Do not use this form to submit sequencing ready samples.
  2. There are different batch sizes depending on library prep, please see our price card or the table above for details.
  3. Researchers from within the CRUK-CI are strongly encouraged to attend an experimental design meeting prior to collecting samples for library prep submission.
  4. Contact CRIExperimentalDesign@cruk.cam.ac.uk to book a free experimental design session.
  5. If you have any questions, contact the Genomics Helpdesk.
CRUK Cambridge Institute, Genomics Core, Li Ka Shing Centre, Robinson Way, Cambridge, CB2 0RE.
(01223) 769 833.
genomics-helpdesk@cruk.cam.ac.uk