30 Cloning Genes of Arabidopsis thaliana by Chromosome Walking

Chromosome walking is a versatile technique applicable to cloning virtually any gene of interest identifiable by mutations1 . The prmciple of this approach consists of using a molecular probe (for instance, a restriction fragment length polymorphism [RFLP] marker) mapping near the target locus to identify genomic clones, particularly those constructed m high capacity vectors The extremities of the cloned insert are recovered and are used m turn as new probes to identify overlappmg clones extending towards the gene. Repeated rounds (or steps) of such screening would eventually yield an overlapping set of clones (contig) bridgmg the mitral RFLP marker and the gene of interestArabidopsis thaliana is particularly suited for isolating specific genes by this approach because of the small size of the haploid content of its genome and the paucity of repetitive DNA 2