Sulfolobus Mutants, Generated via PCR Products, Which Lack
Putative Enzymes of UV Photoproduct Repair
Figure 1
PCR analysis of disruption mutants LR10 and LR12. (a) Outline of the gene-disruption scheme. A plasmid-borne pyrE gene served as template for PCR, which attached S. acidocaldarius chromosomal sequences (gray regions at 5′ ends of the primers) to the ends of this selectable cassette. After the resulting linear DNA was introduced into S. acidocaldarius cells, homologous recombination (indicated by dashed lines) replaced the targeted gene with the cassette. (b) Confirmation of mutant strain genotypes. PCR reactions used genomic DNAs of wild-type S. acidocaldarius (lanes 1, 3, 5, and 7), Saci_1227 disruptant LR10 (lanes 2 and 4), or Saci_1096 disruptant LR12 (lanes 6 and 8) with the following primers: Saci1227 (whole-gene), lanes 1 and 2; Saci1227 internal, lanes 3 and 4; NXSaci1096 (whole-gene), lanes 5 and 6; NXSaci1096 internal, lanes 7 and 8 (see Table 1 for primer data). MW: Molecular weight marker (λ DNA digested with BstEII); fragment lengths (in bp) shown in the right-hand margin.
