Customer can send several kind of material for macroarray screening:
- PCR product (at a minimal concentration of 50 ng/µL ),
- PCR primers, PCR conditions and genomic DNA, in order to amplify the sequence of interest in our laboratory
- Plasmides carrying the required sequence.
Probes are labelled in CNRGV either by:
- random priming labelling method : for PCR products (incorporation of alpha-P33-dCTP)
- or 5' labelling kinase method for oligonucleotides (incorporation of gamma-P33-dATP)
Each probe is purified on a Sephadex column. After hybridization, macroarrays are scanned using a phospho-imager.
Qualitative analysis :
We identify the positive clones using High Density Filter Reader (version 3) software. Based on measures of percentage of black pixels per surface unit, this software allows the identification of positive clones on the hybridized filters:
Each positive clone is finally confirmed by PCR.