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. 2015 Dec 23;6:1170. doi: 10.3389/fpls.2015.01170

FIGURE 1.

FIGURE 1

Chloroplast transformation vector and analysis of transgene integration. (A) Chloroplast genome (WT cpDNA) physical map at the insertion site (dotted lines) showing the left and right flanking recombinogenic regions (LFR and RFR). (B) Vector pBSW-utr/BLSVP8d contains VP8d from BRV strain C486 sequence fused to Brucella spp. lumazine synthase (BLS) under the 5′ untranslated sequence and the promoter of the psbA gene (5′psbA). The LFR includes the 3′ region of rrn16, and the RFR contains the full trnI and the 5′region of trnA. The aadA sequence is under the control of the Prrn promoter. rrn23: sequence coding the 23S rRNA (Wirth et al., 2006). trnI/A: probe used in Southern blot assays. Arrows indicate the location of the forward (Cl Fw) and reverse (Cl Rev) primers used for the PCR analysis. (C) PCR analysis to confirm integration of the recombinant vector into the wild-type plastome. WT: wild-type tobacco plant. The plasmid used for plant transformation was included as a positive control (+). The position of Lambda BstEII marker is indicated on the left. (D) Southern blot using trnI/A probe to confirm integration and homoplasmy (wild-type plastome: 6.4 kbp, transformed plastome: 8.8 kbp). The position of 1-kb DNA marker is indicated on the left. (A-C) independent BLSVP8d transplastomic lines.