METHOD FOR THE RANDOM DIVERSIFICATION OF A GENETIC SEQUENCE WHILE PRESERVING THE IDENTITY OF SOME INNER SEGMENTS OF SAID GENETIC SEQUENCE

PCR method for randomly mutating a nucleotide sequence S, bounded to its extremities 5' and 3' by two segments F1 and F2, by retaining the identity of an internal segment (D) of the sequence, comprises conducting a PCR-error over the entire sequence using primers; selecting the amplification products; conducting a high-fidelity PCR using primer pairs respectively corresponding to at least F1 and D, and F2 and D to amplify the regions F1-D and D-F2 of mutated S in the conducting step, in which D is retained its identity; conducting a high-fidelity PCR; and purifying the PCR products. PCR method for randomly mutating a nucleotide sequence S, bounded to its extremities 5' and 3' by two segments F1 and F2, by retaining the identity of at least an internal segment D of the nucleotide sequence, comprises conducting a PCR-error over the entire sequence S using primers corresponding to the segments F1 and F2 in which the sequence S is randomly mutated along its entire length; selecting the obtained amplification products; conducting a high-fidelity PCR from the amplification products selected in selecting step by using primer pairs respectively corresponding to at least F1 and D, and F2 and D to amplify the regions F1-D and D-F2 of mutated S in the conducting step, in which the internal segment D is retained its identity; conducting a high-fidelity PCR from the amplification products obtained in conducting step by using primers corresponding to F1 and F2; and purifying the obtained PCR products of which the size corresponds to the nucleotide sequence S.