Oligonucleotide-directed mutagenesis: a sequence-based screening.
A procedure to screen for mutant clones obtained by oligonucleotide-directed mutagenesis has been developed. It is based on the preparation of phage containing supernatants from a number (100 or more) of randomly chosen mutagenized M13 plaques. Aliquots from these supernatants are mixed to obtain pools, each containing 10 phages. Heterogeneous single-stranded DNA is prepared from these pools and used as template in a "single letter" sequence according to the dideoxy chain terminator method. Thus, the pool(s) containing the mutated sequence and the mutated sequence itself is identified by sequencing the single-stranded DNAs of the 10 phages present in the selected pool.
Duilio
A
A
Istituto di Scienze Biochimiche, II Facoltà di Medicina e Chirurgia, Università degli Studi di Napoli, Naples, Italy.
Cimino
F
F
Russo
T
T
D000818:Animals; D001483:Base Sequence; D002999:Clone Cells / Q000502:physiology; D003062:Codon; D009154:Mutation / Q000187:drug effects*; D009841:Oligonucleotides / Q000494:pharmacology*; D012361:RNA, Transfer, Pro / Q000235:genetics
D003062:Codon; D009841:Oligonucleotides; D012361:RNA, Transfer, Pro