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DNA SEQUENCING SERVICE

The Centre of Plant Structural and Functional Genomics is equipped with the Applied Biosystems 3730xl 96-capillary Genetic Analyser dedicated to DNA sequencing. We offer DNA sequencing service with a turnaround results within 10 working days at maximum. All our work is performed according to standard operating procedures and to exacting quality control levels.

Please, check the detailed instructions on sending your samples for sequencing and pricing information below.

Contact our staff, should you need any further information.

Results are sent by email in the form of .ab1 files (electropherograms), supplemented with .fas files (text), if required. These files can be viewed using e.g. Chromas or Bioedit (PC platform), which are freely available.

We currently provide two levels of DNA sequencing:

  1. Capillary electrophoresis of customer´s samples.

  2. In house preparation of sequencing reaction.

INSTRUCTIONS FOR SAMPLE PREPARATION

  

1. Capillary electrophoresis of samples that were prepared by the customer using BigDye chemistry

NOTE!: We only accept samples prepared using version 3.1 BigDye chemistry for analysis.

To obtain a high quality data, it is crucial to remove any unincorporated dyes form your sample in the post-reaction clean up. We reccommend to apply either Ethanol precipitation method, Sephadex column purification or Agencourt CleanSEQ kit (Immunotech). Dissolve the samples in deionized sterile water or deionized formamide, according to the purification method used.

In case of inadequate post-reaction clean up, we cannot guarantee the quality of the data.

The minimal acceptable amount of the purified sample to be sent is 10 μl.

Before shipping your samples, please, ensure that the labelling is clear and persistent and the sealing is tight enough to prevent evaporation.

Do not forget to accompany your samples with a printed version of the electronic Sequencing Request Form and Data Sheet. (Supply these documents in electronic form as well!)

  

2. In-house preparation of sequencing reaction (incl. post-reaction clean up) using customer´s DNA templates.

The receipt-confirmation email will be sent to you upon the samples arrival. The sequencing reaction is prepared using BigDye v3.1 according to our optimized protocol. Post-reaction purification is done by Agencourt CleanSEQ kit (Immunotech) according to our standardized sample processing protocol.

Before sending your samples, please, carefully follow instructions below:

(!!! It is the customer's responsibility to provide sufficient amount and appropriate quality of the template and primers)

  1. Mix your purified DNA template in one tube with corresponding primer. We reccomend to use 96-well plates. In case of small number of samples, 0.2 or 0.5 ml tubes are acceptable.
  2. If you are using one of the universal sequencing primers (e.g. M13, T7, T3, SP6, contact our staff for detailed list), provide purified DNA  template only.
  3. Allways make sure that your samples are clearly labelled and appropriate sealing of the plate or tubes is ensured! (we reccommend to use parafilm to prevent evaporation from the tubes).
  4. Send appropriate amount of the template and primer, according to the table:
Template length DNA concentration
PCR product 200-1000 1-4 ng/μl
PCR product 1000-2000 2-8 ng/μl
PCR product over 2000 8-20 ng/μl
Plasmid 20-40 ng/μl
BAC 100-200 ng/μl

The final sample volume must be min. 15 μl.  The final primer concentration must be 2 μM.

NOTE!: In order to streamline the processing of large sample numbers we make no adjustments to concentrations at this end, so we would ask that you ensure your mixture contains the recommended amounts of template and primer.

4.  Before shipment, accompany your samples with a printed and signed version of the electronic Sequencing Request Form and Data Sheet. (Supply these documents in electronic form as well!)

   

PRICING INFORMATION

Commercial customers:
We provide Includes You provide Turnaround results Price
DNA sequencing and basic sequence analysis Preparation of sequencing reaction, post-reaction cleanup, capillary electrophoresis, basic data processing Sequencing primers and DNA template for sequencing prepared as described in Instructions 5-10 working days *

200 CZK (excl. VAT)
Capillary electrophoresis Electrophoretic separation of sample prepared by customer using ABI BigDye chemistry

Purified products of sequencing reaction (see Instructions)

 2-3 working days * 30 CZK (exl. VAT)
    * Please note, that in case of small orders (less than 24 samples), the results delivery time may be prolonged. Contact our staff for detailed information on your specific order.                                                                                                                                                                                                                                                                                                                                                                                                                                                                                  

                                                                                                                            

CONTACTS

Institute of Experimental Botany, AS CR, v.v.i.

Laboratory of Molecular Cytogenetics and Cytometry

Šlechtitelů 31

783 71 Olomouc

Czech Republic

Contact person:

Ing. Marie Seifertová

seifertovamatueb [dot] cas [dot] cz

Tel.: +420 585 238 729

Mgr. Miroslav Valárik, Ph.D.

valarikatueb [dot] cas [dot] cz

Tel.: +420 585 238 714

DOWNLOADS

DATA SHEET
SEQUENCING REQUEST FORM