Towards a Systems Approach in the Genetic Analysis of Archaea: Accelerating Mutant Construction and Phenotypic Analysis in Haloferax volcanii

Figure 2

tRNA analysis. Gene deletions of (a) HVO_1594, (b) HVO_1717 and (c) HVO_1716 were verified by locus-specific PCR using primer pairs as indicated in Supplementary Table 3 designed to anneal outside and within the gene. Predicted amplicon sizes are indicated above each band. Analysis of LC-MS/MS tRNA extracted from H. volcanii strains showing the presence or absence of the m⁵C peak on the UV trace at 254 nm for H26 (d) and the corresponding extraction ion chromatogram for the H26 and ΔHVO_1594 (e). Fragmentation analyses of m⁵C are shown as insets. LC-MS/MS analyses of tRNA extracted from (f) H26, (g) ΔHVO_1717, and (h) ΔHVO_1716 indicate the presence and absence of G⁺. The peaks corresponding to m²₂G and t⁶A are shown for comparison. Fragmentation analyses of G⁺ are shown as insets.

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