Figure 8.

(A) Gel electrophoresis of HaeIII of digested PCR-amplified alkaline-lysis miniprep DNA. A 286-bp fragment of pGBD-MSH2 and pGBD-msh2-M707I DNA produced by directional cloning and isolated from MV1190 transformants by alkaline lysis was amplified by PCR, as described in Materials and Methods. Amplified products were digested with HaeIII, and digest products were fractionated on a 3% agarose + 0.25 μg/ml EtBr gel. Lane 1, 100-bp ladder; lane 2, pMSH2; lane 3, pmsh2-M707I (previously verified by dideoxynucleotide sequencing); lanes 4–9, plasmid DNA from transformants 1–6. Relevant fragment sizes are indicated in the figure. (B) Gel electrophoresis of StyI/KpnI-digested alkaline-lysis miniprep DNA. 1 μg each of pMSH2 (lane 2), pGBD-MSH2 (lane 3), and 6 alkaline lysis miniprep DNA samples (lanes 4–9) were digested with StyI and KpnI, and digest products were fractionated on a 1% agarose + 0.25 μg/ml EtBr gel. DNA was visualized by UV illumination. StyI cleaves once within pMSH2 and once within pGBD-MSH2; KpnI cleaves three times within pMSH2 and once within pGBD-MSH2. Lane 1, 1-kb ladder. Vector (pGBD-MSH2 lacking the smaller StyI-KpnI fragment) and insert (fragment containing the mutagenized region) DNA sizes are indicated in the figure. (Figure and legend courtesy of Ruth Tennen, Princeton University. Reproduced with permission.)