Figure 5: (A ) Illustration of the plastome locus
downstream of psbA prior to insertion of the mVenCP andcodA-aadA cassettes (strain Luc2), as the intermediate
recombinant locus (R1), and following loop-out of the codA-aadAmarker (R2). The locations of the primers (P1 – P4) used in a
four-primer PCR analysis of transformant lines are indicated together
with the expected product sizes. (B ) PCR conformation of
homoplasmy at the R2 stage for four independent Luc2:Ven1 transformant
lines. The Luc2 control showed the expected WT bands at 2.5 kb, whereas
all four transformants showed the expected R2 PCR product at 1.5 kb.
(C ) Spot tests of mid-log cultures, and 1:10 and 1:100
dilutions, of WT, Luc2 and Luc2:Ven1 on TAP plates containing: no
antibiotic, Spc or 5-FC. (D ) Fluorescence analysis of Luc2:Ven1
compare to WT. Fluorescence measurements were taken on
OD750 normalized mid-log cultures as three replicates
(3n). (E ) Microplate-based relative luminescence analysis of
WT, Luc2 and Luc2:Ven1. Luminescence measurements were taken on
OD750 normalized mid-log cultures as three replicates
(3n).