Figure 4: Schematic representation of the FLAEM transformation strategy used to create a markerless deletion mutant.

The general FLAEM method is represented here where Wild-type C. trachomatis is sequentially transformed and serially passaged to generate a markerless deletion mutant. Transformation steps are represented by the addition of small arrows, and the loss of genetic elements is represented by the subtraction of small arrows. C. trachomatis intermediates (circles) are represented with antibiotic sensitivities (penicillin-resistant or penicillin-sensitive = PenG^r or PenG^s; spectinomycin-resistant or spectinomycin-sensitive = Spec^r or Spec^s) and fluorescence-reporting qualities (green = gfp +, red = mCherry +, gray = no fluorescence). Schematic representations of the gene locus at each step are shown below. This figure has been modified from Keb et al.¹².