High throughput synthesis of long oligonucleotides


  HEADGOUYETTE Catherine / oligos@pasteur.fr
  MEMBERS


  Annual Report

Inside this platform, we synthesize long

oligonucleotides for platforms PF2 (DNA microarrays),

and PF1 (Genomics) projects, most of the time

selected by steering committees and for all the units

of the Institut Pasteur which sometimes, prefer

to appeal to our ability (specially in quality

controls that we make on our production) instead of

buying the oligos from private companies.

The methodology we use, is based on the phosphoramidite chemistry employed by all the automatic synthesizers and this should not change in the coming years as it is very efficient (given its rapidity and the very good yields in coupling reactions).

We are able to make about 570 oligos (70mers) a week for microarrays purposes. The platform is also making modified oligonucleotides for other purposes (HPLC purified molecular beacons, siRNA, 5’- or 3’- modified oligos by fluorophores or amino-groups).

Since its creation, in January 2003, the platform is associated with the Genopoles National Network and co-operates with many research teams inside this Institute such as Legionnella pneumophila, Entamoeba histolytica, Plasmodium falciparum, or by taking part in international programs on big projects such as: Candida glabrata.

Quality controls are based upon capillary electrophoresis, high performance liquid chromatography, whatever is the length of the oligos and for oligos up to 40-50 bases, a third control is based on mass spectrometry after matrix laser desorption.

We synthesized:

.5’-amino-oligos covalently fixed on an aldehyde modified glass support, to develop microarrays dedicated to the analysis of expression profile of micro RNAs;

.5’-Cy5 or 5’-Cy3-oligos prepared for making diagnostic microarrays in view of identification of different Plasmodium stems;

.Several milligrams of short oligos for spectroscopic studies: cristallography, X-ray diffraction and NMR (publications with Pr Subirana at the University of Catalunya, Spain) and for the same purpose RNA sequences for Prof Ghomi (UMR CNRS 7033, Paris 13 University);

.Molecular beacons, oligos with a 5’-fluorophore group and a quencher at the 3’ end to visualize the mRNA transport in the cell;

.And, many siRNA: study of the extinction by siRNA of the multiplication of RNA-positive viruses of medical interest (hepatite C virus , enteroviruses).



  Web Site

More informations on our web site


  Publications

Publications 2006 of the unit on Pasteur's references database




Activity Reports 2006 - Institut Pasteur
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