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. 2020 Jan 22;9:468. doi: 10.3389/fcimb.2019.00468

Figure 2.

Figure 2

Generation of the L. braziliensis co-expressing eGFP and LUC. (A) Agarose gel electrophoresis of pIRSATeGFP_LUC following SwaI digestion (lane 1), 1Kb marker (lane 2), Undigested circular plasmid (lane 3). (B) Fluorescent microscopy of transgenic L. braziliensis clones. (C) Agarose gel electrophoresis of PCR product amplified using primers complementary to the Nourseothricin resistance marker (SAT) present in the of integration cassette and complimentary to the rDNA locus of the L. braziliensis genome (outside the cassette), indicative of eGFP-LUC integration, SSU locus (18s gene). PCR was performed using genomic DNA purified from WT Lb (lane 1) or from transgenic Lb (clones 6 and 7) (lane 3 and 4, respectively). (D) Agarose gel electrophoresis of PCR product amplified using primers complimentary to the ITS-1 locus using genomic DNA purified from WT-Lb (lane 2) or eGFP-LUC Lb (transfected clone 7) (lane 3) (Left panel). HaeIII digestion of ITS-1 PCR product. WT-Lb (lane 2) or transgenic Lb (transfected clone 7) (lane 3) (right panel).