(classical) genetics causality phenotype DNA ‘reverse’ genetics

T C

The ‘gapped duplex’ method of in vitro mutagenesis

Gapped duplex method with phenotypic selection

Ceftazidime

S K G mutant TEM bla 5´ p GAT AAA TCT GGA GCC TCC AAG GGT GGG TCT CGC GG 3´ -----gat aaa tct gga gcc GGT GAG Cgt ggg tct cgc gg G E R wild-type TEM bla

Creating or removing a restriction recognition site as an aid in screening for site-specific mutants Target : replace Trp by Phe Screening aid : MnlI cleavage Mismatch primer : …GCCCTGGGCTTCGGTGGCA…

Codewoord tabel voor ‘reverse translation’ Leu = 1T2 Arg = 3G4 Ser = 567

Ceftazidime is a third-generation cephalosporin antibiotic. Like other third-generation cephalosporins, it has broad spectrum activity against Gram-positive and Gram-negative bacteria. Unlike most third-generation agents, it is active against Pseudomonas aeruginosa, however it has weaker activity against Gram-positive microorganisms and is not used for such infections.cephalosporinantibioticGram-positiveGram-negativebacteriaPseudomonas aeruginosa Ceftazidime is a semisynthetic, broad-spectrum, beta-lactam antibiotic for parenteral administration. Ceftazidime is bactericidal in action exerting its effect by inhibition of enzymes responsible for cell-wall synthesis. A wide range of gram-negative organisms is susceptible to ceftazidime in vitro, including strains resistant to gentamicin and other aminoglycosides. In addition, ceftazidime has been shown to be active against gram-positive organisms. It is highly stable to most clinically important beta-lactamases, plasmid or chromosomal, which are produced by both gram-negative and gram-positive organisms and, consequently, is active against many strains resistant to ampicillin and other cephalosporins. Ceftazidime has activity against the gram-negative organisms Pseudomonas and Enterobacteriaceae. Its activity against Pseudomonas is a distinguishing feature of ceftazidime among the cephalosporins.