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RNA library preparation

RNA-seq

RNA-seq is a used for sequencing of transcriptomes. High quality, DNAse-treated (critical!) total-RNA is used as starting material for library preparation.  All libraries are indexed and typically paired-end compatible. The amount of starting material varies on the sample preparation method selected. Sample preparation method is selected together with you. Options:

  • non-strand-specific method
  • strand-specific
  • low RNA amounts are supported.

Either ribosomal-RNA depletion or polyA-RNA-isolation can be used to discard ribosomal-RNA. In library preparation methods with small amount of starting material the ribosomal RNA may not be depleted at all.  Typically 2-4 mammalian transcriptomes are run in one HiSeq lane.

Customer provides

  •  1-5 µg high quality, DNAse-treated total-RNA or 50-250ng of mRNA
  •  Sample information sheet including the sample IDs and other required info
  •  Customer will apply for FIMM user account for data delivery

Sequencing unit delivers:

  • Sample quality control
    - Agilent Bioanalyzer (quality and size distribution)
    - Qubit RNA (quantity)
    - Qubit DNA (quantitation of DNA contaminants)
  • Library preparation with selected protocol
  • Size selection of the sequencing library
  • Quantitation and fragment size distribution of the library
    - Bioanalyzer HS or Caliper GX HiSens analysis

SmallRNA /microRNA library preparation

smallRNAseq is used for sequencing of small RNA fraction of the transcriptome (<200nt). High quality, DNAse-treated total-RNA containing small RNA-fraction is used as starting material for sample preparation to create Illumina compatible indexed library.

Customer provides

    1ug high quality, DNAse-treated total-RNA
    Sample information sheet including the sample IDs and other required info
    customer will apply for FIMM user account for data delivery

Sequencing unit delivers:

  •  Sample quality control
    - Agilent Bioanalyzer (quality and size distribution)
    - Qubit RNA (quantity)
    - Qubit DNA (quantitation of DNA contaminants)
  • Preparation of smallRNA sequencing libraries
  • Size selection of smallRNAs of interest (typically miRNAs)
    - Caliper XT instrument
  • Quantitation and fragment size distribution of the library
    - Bioanalyzer HS or Caliper GX HiSens analysis
Last updated: 12.09.2017 - 15:54