Process for in vitro creation of recombinant polynucleotide sequences by oriented ligation

Process according to any one of the preceding claims, wherein a preferably thermostable ligase that is active at high temperature is used at step (d).

Process according to any one of the preceding claims, wherein the initial bank of polynucleotide sequences is produced from a wild gene by successive steps of controlled mutagenesis, by error prone PCR, by random chemical mutagenesis, by in vivo random mutagenesis, or by combining genes of near or distinct families within the same species or from different species so as to make available a variety of polynucleotide sequences in the initial bank.

Process according to any one of the preceding claims, wherein the initial bank is fragmented into n fragments at step (a), n being greater than or equal to 3.

Process according to any one of the preceding claims, wherein besides the matrix, oligonucleotides of varying length, single- or double-stranded, are added at step (b) or (c).

Process according to any one of the preceding claims, wherein, before step (e), the recombinant polynucleotide sequences are separated from the assembly matrix thanks to a marker present on the assembly matrix or on the recombinant polynucleotide sequences.

Process according to any one of the preceding claims, wherein the recombinant polynucleotide sequences obtained at step (d) and optionally cloned are used by any appropriate means to select the recombinant polynucleotide sequences or the clones offering advantageous characteristics compared with corresponding characteristics of reference sequences.