Fig. 1.

Transposons used for insertion mutagenesis. Transposons ISphoA/ hah (4.83 kbp) and ISlacZ/hah (6.16 kbp) are derived from the IS50L element of transposon Tn5 and generate alkaline phosphatase (′phoA) or β-galactosidase (′lacZ) translational gene fusions if appropriately inserted in a target gene. An outward-facing neomycin phosphotransferase promoter is expected to reduce polar effects on downstream gene expression for appropriately oriented insertions. Cre-mediated recombination excises sequences situated between the loxP sites in each transposon, leaving a 63-codon insertion that encodes an influenza-hemagglutinin epitope and a hexahistidine metal-affinity purification tag (together referred to as ”hah,” see ref. 10). ′phoA, alkaline phosphatase gene; ′lacZ, β-galactosidase gene; tet, tetracycline resistance determinant; loxP, Cre recognition sequence; P, neomycin phosphotransferase promoter.