Technical Specifications

Work Flow

  • Sample QC Incoming extracted DNA and RNA is analysed to determine the quality of the fragments and the quantity of the sample. If the sample falls below the recommended quality or quantity for the library preparation, you will be contacted to make a decision on how to proceed.

  • Library Preparation: The selection of the library preparation is made depending on the type of sequencing being undertaken and the specific sample submitted. Supplier protocols are followed for the library preparation to ensure the quality of the library for sequencing.

  • Library Validation: For routine library validation, another rigorous quality control is performed to ascertain the library fragment size, yield and concentration.

  • Cluster Generation: During cluster generation, the library hybridises to the flow cell and a series of bridge amplifications occur.

  • Sequencing by Synthesis: This process involves the addition of fluorescently labelled reversible-terminator nucleotides which compete to bind the strand tethered to the flow cell. Once the correct base binds, laser excitation is used to emit a wavelength, and this specific wavelength is detected by a high resolution camera. Once this has been detected, the fluorescent and terminator sequences of the nucleotide are cleaved, allowing the addition of a new fluorescent nucleotide. This occurs in a “massively parallel” fashion to obtain large amounts of sequencing data.

Sample requirements

Each library preparation kit has differing minimum sample quantity and concentration values, depending on the application, input quality and technology. We require more product than the minimum specified by the manufacturer as we require excess DNA/RNA to perform our quality control steps.

If your samples do not meet the minimum sample requirements, we will contact you to discuss the best progression path for your project.

Please contact us for advice on which options may be suitable for your samples.

Deliverables

Once all analysis has been completed, all data is returned to the customer via secure FTP site download or external hard drive. This will include:

  • All intermediate files generated during analysis. Retention of this data by the customer allows further experiments to be carried out in the future, without needing to re-sequence samples

  • Final (filtered) output data, in FASTA or FASTQ format. FASTQ is a modification of the popular FASTA format, with quality scores assigned to each base-call.

  • Data from all bioinformatics analysis that have been performed. File formats will depend on the particular type of analysis.

  • Lab QC report detailing the integrity of the samples that were used for sequencing.

  • Bioinformatics pipeline report, explaining quality control metrics used to assess the quality of the data, and details of all bioinformatics analyses that have been performed, explaining the methods used and results obtained.