In-gel DNA radiolabelling and two-dimensional pulsed field gel electrophoresis procedures suitable for fingerprinting and mapping small eukaryotic genomes

Jean-François Brugère (1) , Emmanuel Cornillot (2) , Guy Méténier (2) , Christian Vivarès (2)

1 CIDAM - Conception, Ingénierie et Développement de l'Aliment et du Médicament
2 LMGE - Laboratoire Microorganismes : Génome et Environnement

A simple method for complete genome radiolabelling is described, involving long-wave UV exposure of agarose-embedded chromosomal DNA and [α-32 P]dCTP incorporation mediated by the Klenow fragment. Experiments on the budding yeast genome show that the labelling procedure can be coupled with two new two-dimensional pulsed field gel electrophoresis (2D-PFGE) protocols of genome analysis: (i) the KARD (karyotype and restriction display)-PFGE which provides a complete view of the fragments resulting from a single restriction of the whole genome and (ii) the DDIC (double digestion of isolated chromosome)-PFGE which is the eukaryotic counterpart of complete/complete 2D-PFGE in bacterial genomics.

Domaines: Alimentation et Nutrition; Génomique, Transcriptomique et Protéomique [q-bio.GN]; Interactions cellulaires [q-bio.CB]; Systématique, phylogénie et taxonomie; Evolution [q-bio.PE]; Biotechnologies; Santé; Ecosystèmes; Interactions entre organismes; Hépatologie et Gastroentérologie; Cardiologie et système cardiovasculaire; Bactériologie; Biochimie [q-bio.BM]; Biologie moléculaire; Organisation et fonctions cellulaires [q-bio.SC]

Jean-François Brugère, Emmanuel Cornillot, Guy Méténier, Christian Vivarès. In-gel DNA radiolabelling and two-dimensional pulsed field gel electrophoresis procedures suitable for fingerprinting and mapping small eukaryotic genomes. Nucleic Acids Research, 2000, 28 (10), pp.48e - 48. ⟨10.1093/nar/28.10.e48⟩. ⟨hal-01612762⟩