Figure 2.

A system for constructing chromosomal deletion in M. jannaschii employing the Hmg-CoA reductase gene (hmgA) as a selectable marker. Construction of a M. jannaschii BM10 (Δfsr::P_sla.hmgA) strain. (A) Replacement of fsr via double cross-over recombination between the upstream (UP) and downstream (DN) regions of fsr (locus tag number, mj_0870) and cloned homologous elements in a linearized form of the suicide plasmid, pDS210 (Figure 1A). (B) Characterization of M. jannaschii BM10 genotype by the use of PCR analysis. PCR was performed on isolated chromosomal DNA of M. jannaschii BM10 employing primers 1 and 2 as shown in A; primer sequences appear in Supplementary Table S1. The genomic regions targeted for PCR amplification are shown with bold double-sided arrows in (A). Sizes of DNA markers (M) and the PCR amplicons are shown next to the respective DNA bands. (C) Growth phenotypes of M. jannaschii wild-type and BM10 strains on Gelrite plates under three conditions: without selection, under mevinolin (10 μM) selection, and in the presence of 10 mM sulfite. Details of some of the abbreviations and pDS210 appear in the legend of Figure 1.