Figure 2.

Designer transposons used in a marker-less approach (A) and a marker-driven approach (B). Asterisk denotes a 19-bp mosaic end recognized by Tn5 transposon. In (A), a query fragment can be generated using polymerase chain reaction or prepared as a synthetic DNA fragment. In (B), a query ORF is cloned downstream of the pac puromycin resistance marker. The start codon of the query ORF immediately follows the stop codon of the pac gene (see Supplementary Figure 2). This ORF can be amplified from a template or prepared as a synthetic DNA block. The tuf promoter that drives the expression of the pac gene also drives the expression of the query ORF. T denotes transcriptional terminator.