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site-directed munogenesis is a common method to introduce specific alterations into a gene of interest once the gene has been cloned into a plasmid vector synthetic oligonucleotides with the desired mutations can be used to introduce the mutation into the gene oligonucleotides can be designed to introduce point mutations insertions or deletions into the gene sequence here a mutation will be introduced into section b of the gene the template dna must be single stranded circular dna as an m13 a filamentous bacteriophage the mutant oligonucleotide here a deletion mutation is annealed to the gene of interest and extended using a dna polymerase to generate a mutant copy the mutant oligonucleotide contains complementary sequences to the a and c regions of the gene of interest with a deletion in region b once the oligonucleotide is annealed dna polymerase dna ligase datp dgtp dttp alpha s dctp and magnesium are added to the reaction mixture this allows for the extension of the gene product us
