Abstract: The invention concerns the introduction of predetermined genetic changes in target genes of a living cell by introducing an oligodeoxynucleotide encoding the predetermined change. The oligodeoxynucleotides are effective in mammalian, avian, plant and bacterial cells. Specific end modifications that greatly increase the effectiveness of the oligodeoxynucleotides in bacteria are described. Surprisingly, unmodified oligodeoxynucleotides can be as effective in mammaliancells, including in vivo hepatocytes, as the modified nucleotides and can be as effective or more effective than chimeric oligonucleotides that consist of a mixture of deoxynucleotides and 2′-O-methyl ribonucleotides.

Abstract: The invention is based on the discovery that recombinagenic oligonucleobases are active in prokaryotic cells that contain a strand transfer activity (RecA) and mismatch repair activity (MutS). Using this system a type of Duplex Mutational Vector termed a Heteroduplex Mutational Vector, was shown to be more active in prokaryotic cells than the types of mutational vectors heretofore tested. Further improvements in activity were obtained by replacing the tetrathymidine linker by a nuclease resistant oligonucleotide, such as tetra-2′-O-methyl-uridine, to link the two strands of the recombinagenic oligonucleobase and removing the DNA-containing intervening segment. The claims concern Duplex Mutational Vectors that contain the above improvements. In an alternative embodiment the claims concern the use of Duplex Mutational Vectors in prokaryotic cells.

Abstract: The invention is based on the discovery that Duplex Mutational Vectors are active in prokaryotic cells that contain a strand transfer activity (RecA) and mismatch repair activity (MutS). Using this system a type of Duplex Mutational Vector, termed a Non-Chimeric Mutational Vector, having no RNA:DNA hybrid-duplex, was shown to be active in prokaryotic cells if protected from 3' exonuclease attack. Such protection can be conferred by replacement of a tetrathymidine linker by a nuclease resistant oligonucleotide, such as tetra-2'-O-methyl-uridine, to link the two strands of the recombinagenic oligonucleobase. The claims concern 3'-exonuclease protected Non-Chimeric Mutational Vectors.