From a short amino acidic sequence to the complete gene

A useful strategy directed to the isolation of a required gene with a high GC content is reported. Using a degenerate oligonucleotide probe, deduced f rom the amino terminus of a protein, it is possible to obtain a fragment of DNA containing its encoding gene by PCR amplification. Furthermore, the cl oning of a desired gene can be accomplished in two steps by using an oligon ucleotide deduced (i) from an internal sequence, (ii) from a consensus sequ ence, or (iii) from a DNA sequence adjacent to a disrupting element (transp oson, insertion sequence, cassette). This method, which could be applied to a bacteriophage, plasmid, or cosmid genomic library, has been successfully used for cloning several genes from different biological systems. (C) 2000 Academic Press.