Patent ID: 7790374

Claim:
A process for the mutagenesis of a double-stranded polynucleotide sequence (master sequence) of n base-pairs having a (+)-strand and a complementary (−)-strand comprising the steps (i) creating a collection of single-stranded fragments of the (+)-strand of the master sequence wherein all members of the collection have the same 5′-terminus and have a deletion in the 3′-terminus such that the collection represents (+)-strands with a length of n−1, n−2, n−3, . . . nucleotides; (ii) introducing at least one universal or degenerate nucleotide at the 3′-terminus of the (+)-strands produced in step (i); (iii) elongating the (+)-strands produced in step (ii) to the full length of the master sequence using the (−)-strand or fragments thereof of the master sequence as a template strand for the elongation; and (iv) synthesizing a (−)-strand by using the (+)-strand produced in step (iii) as a template strand thereby effecting mutations in the (−)-strand at the positions of the previous universal or degenerate nucleotides compared to the master sequence.