PRINS Reaction

1. Primers: Alpha-satellite primers able to discriminate between chromosomes were produced for the human (12,13). In the present study, we used the followed primers, which specific for centromeres of human chromosomes 1 and 7, respectively:

1c: 5'ATTCCATTAGATGATGACCCCTTTCAT3' 7c: 5'AGCGATTTGAGGACAATTGC3'

2. Taqpolymerase and specific buffer, including MgCl2 (Qbiogen, Illkirch, France).

3. 2'-Deoxyribonucleotide 5'-triphosphate (dNTPS): Ultrapure dNTPs set (each 100 mM Pharmacia Biotech, St Quentin en Yveline, France).

4. Fluorescein-12-2'-deoxyuridine 5'-triphosphate (dUTP; Roche Molecular Biochemicals, Meylan, France (see Table 1).

5. Slides: 76 x 26x 1 mm (SuperFrost LLR2BN from CML, Nemours, France).

6. Thermo cyclers: we used two cyclers alternatively; the Hybaid Omnigen (Hybaid Limited, Teddington, UK) and the Crocodile III, Appligene (Qbiogen).

7. Rubber cement.

10. dNTPs mix (aliquoted by 10 pL and frozen): 120 pL of pure distillated water, 20 pL of each 100 mM dATP, dTCP, dGTP, 20 pL of a mother solution of 2.5 mM 2'-deoxythymidine 5'-triphosphate (dTTP; 1/40 dilution of the original 100 mMdTTP).

11. Washing solution: 4X SSC, Tween-20, pH 7.0-7.2 (200 mL 20X SSC, 5 mL of a 10% aqueous solution realized from Tween-20 and 795 mL of distillated water; see Note 2).

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