Science

Sanger sequencing

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illustration of a method of dna sequencing

The Ramaciotti Centre for Genomics is committed to providing a quality Sanger DNA sequencing service with a fast turnaround time. Our services are flexible and can be customised to meet your needs. All sequencing performed by the Ramaciotti Centre for Genomics is carried out in our ISO/IEC 17025 accredited laboratories.

If the Sanger sequencing service you require is not listed on the service menu below or you need assistance with project design please contact us.

If you are a returning customer, please submit your order through the LabLink portal. 

  • Our Core Prep services are an easy option for researchers who would like to fast track their project or for those who are new to Sanger sequencing.

    Core prep

    You submit a mix of purified PCR product and primer, we perform the sequencing reaction and capillary separation.
    Turnaround time: 3-5 days.*

    Core prep with clean up

    You submit unpurified PCR product and primer, we perform the PCR clean up, sequencing reaction and capillary separation. Clean up options include ExoSAp-IT and magnetic bead clean up (plate submissions only).  
    Turnaround time: 3-5 days.*

    Returning customers

    Please log in to our LabLink portal to submit your samples.

    New customers

    Please download our user guides for detailed instructions on how to use our Sanger sequencing services:

    Data delivery

    Sanger sequencing results will be available in Lablinks, under each Sanger project's Results and Documents tab. The following files are available to download:

    • ABI format chromatogram
    • QUAL file
    • SEQ file

    Quality guarantee

    • We include internal controls (pGem) on all sequencing runs to ensure quality data.
    • All traces are manually inspected before data is released.
    • Free re-run service if results do not meet our quality standards.

     

    *  Working days are Monday – Friday (excluding Public Holidays). Samples must be received by 12 noon to be included in the day's sequencing run.

  • Our User Prep services are useful for researchers who are hands-on and have access to the required sequencing reagents.

    User prep 

    You submit clean, pelleted sequencing product, we perform the capillary electrophoresis. Turnaround time: 1-3 days.*

    User prep plus

    You submit the product of the sequencing reaction, we perform the clean up and the capillary electrophoresis. This service is only available for plate submissions. 
    Turnaround time: 1-3 days.*

    Returning customers

    Please log in to our LabLink portal to submit your samples.

    New customers

    Please download our user guides for detailed instructions on how to use our Sanger sequencing services:

    Data delivery

    Sanger sequencing results will be available in Lablinks, under each Sanger project's Results and Documents tab. The following files are available to download:

    • ABI format chromatogram
    • QUAL file
    • SEQ file

    Quality guarantee

    • We include internal controls (pGem) on all sequencing runs to ensure quality data.
    • All traces are manually inspected before data is released.
    • Free re-run service if results do not meet our quality standards.

    *  Working days are Monday – Friday (excluding Public Holidays). Samples must be received by noon to be included in the day's sequencing run.

  • 1. Sanger sequencing

    Successful Sanger sequencing is achieved by good template-primer complementary and template-primer ratio. The table below shows the recommended quantity of template to use in a sequencing reaction base on the size of the targeted template . The AB3730 DNA Analyzer is a highly sensitive instrument, consequently you may not need as much template as indicated in the table. Input amounts may have to be optimised as over-saturation of signal will cause a reduction in data quality.

    As a starting point we suggest that clients read our Sanger Sequencing Guide. 

    Template Recommended quantity
    PCR - 100 - 200bp
    200 - 500bp
    500 - 1000bp
    1000 - 2000bp
    >2000bp    		 1 - 3 ng
    3 - 10ng
    5 - 20ng
    10 - 40ng
    40 - 100ng
    Single-Stranded DNA 50 - 100ng*
    Double-Stranded DNA (Plasmid DNA) 200 - 500ng*
    Large DNA (e.g. BACs,PACs,YACs,cosmids and fosmids) 0.5 - 1.0ug
    Bacterial genomic DNA 2 - 3ug

    Table referenced from DNA Sequencing by Capillary Electrophoresis by Applied Biosystems.

    2. Sequencing guides

    Below are links to resources for general information on Sanger sequencing and troubleshooting your data:

    Ramaciotti Centre Sanger Sequencing Guide

    DNA Sequencing by Capillary Electrophoresis by Applied Biosystems

    Qiagen Guide to Template Purification and DNA Sequencing

    3. Sequencing clinic

    The Centre runs a sequencing clinic whereby clients can book one on one time with a Sanger sequencing expert. Please contact us for further information or to make an appointment.

    4. Software for viewing and editing chromatograms

    We advise our customers to check the chromatograms manually to ensure correct base calling and genotype calling. There are a large number of programs available, each having its own specific advantage. 

    • Applied Biosystems Sequencing Analysis 5.1.1 for Windows XP and SeqScape (Sequencing Analysis and alignment) from Applied Biosystems.
    • FinchTV software for viewing and editing chromatograms (Macintosh OS X, Windows, Linux). Free access.
    • Geneious is a comprehensive suite of tools for analysis of a range of sequencing data. Trial and student discounts are available.
    • Genemapper 5.0 for Windows 7 is available for Microsatellite, AFLP and RFLP analysis from Applied Biosystems.
    • Sequence Scanner (Sequencing Analysis) and Peak Scanner (Fragment Analysis) Applied Biosystems for PC. Free access.
       

    The Centre does not provide software for analysis of sequencing data but can provide an analysis service if required (conditions and fees apply). Contact us for further information.
  • Service type Submission format Reaction number Price/Reaction (ex GST)
    Core Prep

    Tube

    Plate

    Any

    40 - 96

    $11.50

    $9.50
    Core Prep & ExoSAP-IT

    Tube

    Plate

    Any

    40 - 96

    $13.50

    $11.50
    Core Prep & Bead Clean Up Plate 40 - 96 $13.50
    User Prep

    Tube

    Plate

    Any

    40 - 96

    $7.00

    $5.00
    User Prep Plus Plate 40 - 96 $7.20

    Included in our services:

    • Routine quality checking of all traces.
    • Easy access to your sequencing data.
    • Technical support.
    • Troubleshooting advice.

  • Software for viewing and editing chromatograms

    There are a number of analysis program options available:

    • Applied Biosystems Sequencing Analysis 5.1.1 for Windows XP and SeqScape (Sequencing Analysis and alignment) from Applied Biosystems.
    • Genemapper 3.7 for Windows XP is available for Microsatellite, AFLP and RFLP analysis from Applied Biosystems.

    The Centre does not provide software for analysis of sequencing data but can provide an analysis service if required (conditions and fees apply). Please contact us for further information.

    Other analysis programs (including freeware)

    • Sequence Scanner (Sequencing Analysis) and Peak Scanner (Fragment Analysis) Applied Biosystems for PC.
    • Free EditView software from Applied Biosystems for viewing and editing chromatograms (Macintosh OS 9.x, 1.5 Mbytes).
    • Free FinchTV software for viewing and editing chromatograms (Macintosh OS X, Windows, Linux).
    • Free 4Peaks software for viewing and editing chromatograms (Macintosh OS X).
    • BioEdit – Biological sequence alignment editor for Windows 95/98/NT.
    • Genedoc – a multiple sequence alignment editor, analyser, and shading utility for Windows.
    • STRand – http://www.vgl.ucdavis.edu/STRand (Fragment Analysis) for PC.

    Other helpful links and guides

    • Genbank – sequence submission support and software.
    • BLAST – a set of similarity search programs designed to explore all of the available sequence databases regardless of whether the query is protein or DNA.
    • Entrez – an integrated search and retrieval system that takes information from all of the databases at NCBI.
    • Genographer – The MSU Sequencer Users’ Group, EPSCoR Plant Biotechnology Group, Barley Genetics, Genographer and Barley Production Homepages.
    • The Association of Biomolecular Resource Facilities