Guidelines to sample deposit

  1. Prepare your samples
  2. Fill out the online request form
  3. Print out a copy of the PDF and leave it with the tubes

Samples and primers have to be in 1.5ml tubes:

  • One tube with DNA (2µL/reaction)
  • One tube with primer (2µL /reaction, at 5µM)

If a sample/primer is used more then once, increase the volume in the same tube.

Resuspend your DNA in purified water. Do not resuspend DNA in TE, as EDTA might inhibit the sequencing reaction.

Verify the concentration of plasmid DNA with a spectro. For PCR products, migrate on an agarose gel (after purification) with a molecular weight marker of known concentration and compare.

Required concentrations and quantities

Matrix Concentration Quantity
PCR products
100 - 200 pb
200 - 500 bp
500 - 2000 pb
2000 pb and more

1 ng/µl
5 ng/µl
10 ng/µl
25 ng/µl

2 µl/reaction
minimum 5 µl
Plasmid DNA 200 à 300 ng/µl 2 µl/reaction
minimum 5 µl
BAC 1 µg/µl 5 µl/reaction

Primers

The following primers are available for free :

M13+       5'- TGT AAA ACG ACG GCC AGT -3'
M13-5'- TCA CAC AGG AAA CAG CTA TGA C -3'
SP65'- GAT TTA GGT GAC ACT ATA G -3'
T35'- ATT AAC CCT CAC TAA AGG GA -3'
T7+5'- TAA TAC GAC TCA CTA TAG GG -3'
T7-5'- GCT AGT TAT TGC TCA GCG G -3'

Primer design (free software available: Primer 3, PrimerQuest)

  • Length between 18 and 24 nucleotides.
  • GC content between 40 % and 60 %.
  • Tm between 50°C and 60°C.
  • Avoid regions of repeated nucleotides (ex: AAAAA ou GGGGGG).

Where to deposit/send your samples

Plateforme génomique, IRIC
Université de Montréal
Pavillon Marcelle-Coutu, labo 4462
2950, Chemin de Polytechnique
Montréal, H3T 1J4

If you bring samples in person,
please deposit samples on the 4th floor next to the elevators

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Address if you’re using a delivery service:
Raphaëlle Lambert
Plateforme génomique, IRIC
Université de Montréal
Pavillon Marcelle-Coutu
2950, Chemin de Polytechnique
Montréal, H3T 1J4

The platform is open from Monday to Friday between 9AM and 5PM.

Running samples

  1. Sequencing reactions using Applied Biosystem's BigDye® Terminator v3.1
  2.   25 cycles  
      curly brace  
    96°C 96°C 50°C 60°C 4°C
    4 min15 sec5 sec4 min
  3. Purification by ethanol precipitation
  4. Migration by capillary electrophoresis using an Applied biosystems 3730 automatic sequencer (48 capillaries).

Retrieving your sequences

Each user has a personal access to his sequences on a secured server.

In the « Secured Section », identify yourself with your email and password. Then, the sequences are available in text format (.txt) and in binary chromatogram format (.ab1). We recommend the visualisation of your chromatograms to detect problems that are invisible in the text file (second sequence, contamination, bad quality sequence, etc.).

We don't guarantee good sequencing results as they reflect of the quality of your samples. A good reaction will yield a maximum of 900 to 1000 bp. Purity and concentration of the matrix are the most important factors to obtain a good quality sequence. This is why it is very important that you verify the quality and concentration of your DNA, and the concentration and specificity of your primers before you send it for sequencing. We use a positive control to ensure that the sequencing reaction and the migration go well. A good reaction will yield 1000 bp.

We keep your samples for a maximum of 2 weeks.

If you have any questions, contact us using our contact form