INVESTIGADORES
PETRUCCELLI Silvana
artículos
Título:
Recovery of Tn5-Flanking Bacterial-DNA by Vector-Mediated Walking from the Transposon to the Host Genome
Autor/es:
HOZBOR, DF; OTERO, AJLP; WYNNE, ME; SILVANA PETRUCCELLI; LAGARES, A
Revista:
ANALYTICAL BIOCHEMISTRY
Editorial:
Elsevier
Referencias:
Lugar: Maryland; Año: 1998 vol. 259 p. 286 - 288
ISSN:
0003-2697
Resumen:
Genetic studies to identify bacterial genes related to specific biochemical functions are usually accomplished by classical strategies of random mutagenesis, phenotypic screening to detect mutants of interest, and cloning of the mutagenized DNA fragments (1, 2). Different transposons have been widely used for the initial step of mutagenesis (3) since many of them generate random and relatively stable insertional mutations, which can be later cloned and selected using an antibiotic resistance coded by the transposable element. Tn5 has been particularly useful, since it transposes at a high frequency in a variety of gram-negative bacteria with a low insertional specificity (5818 bp, kanamycin/neomycin (Km/Neo),2 bleomycin (Ble), and streptomycin (Str)) (4). However, cloning of mutagenized DNA is frequently a cumbersome step as a consequence of the low efficiency of the bimolecular