BAC Modification through Serial or Simultaneous Use of CRE/Lox Technology
Figure 5
Specificity of Lox variant site reactions. (a) Vector map of Lox-flanked selection cassettes in a single-copy BAC backbone. Lox site locations indicated in green boxes and Ampicillin cassette indicated in teal box. (b) At top, organization of Lox sites and selection cassette in vector prior to Cre expression. Below are listed the expected products following Cre expression. Primer locations are indicated by arrows. In PCR analyses, the original cassette produces a 360 bp band, while the Cre-mediated products of deletion or inversion produce bands of 240 bp and 171 bp, respectively. (c) Gel electrophoresis of colony PCR analysis of EL350 bacterial bearing the BAC testing vector after induction of Cre expression and plating. PCR-positive bands indicate when a deletion or inversion has taken place (lanes 1, 2, and 3, (left)). PCR-positive bands indicate when unmodified clones and clones with an inversion are present (lanes A, B, and C, (right)). One Kb Plus DNA ladder (Ladder) is displayed at the right of each gel photograph. The molecular weight of relevant bands is indicated next to the gel on the right.
