CHROMOSOME WALKING IN DICTYOSTELIUM - AN APPLICATION OF THE SINGLE SPECIFIC PRIMER PCR

We describe here an application of the polymerase chain reaction that sidesteps the necessity of knowing sequence information at both ends o f a desired fragment of genomic DNA. The methodology is straightforwar d and works efficiently in organisms with relatively simple genomes. W e have used it to walk more than 8 kb in six steps from a 'starter' DN A sequence that was identified in a cDNA library screen. This technolo gy complements existing procedures to walk in either direction along c hromosomes; but because of the multiple levels of selection employed, it may prove to be more effective. Its primary utility will probably l ie in extending sequence information in simple organisms that do not h ave pre-existing cDNA or genomic libraries, or for rapidly isolating g enomic sequences that flank a cDNA clone. (C) 1997 Elsevier Science B. V.