Restriction digests of the clone give the following sizes (kb): EcoRI--4.4; BamHI--4.4; HindIII--4.4; PstI/XbaI--2.9, 1.5. Two rounds of gel purification of the restriction digest generated probe is necessary. Failure to do so often results in high backgrounds (due to flanking regions in the plasmid insert) and false positives with clinical E. coli strains. Recommendation for verification: XbaI; PstI; XbaI/PstI; RsaI; ClaI. The 273bp RsaI fragment is recommended as a hybridization probe for aac(6')-IIb. |
