Regeneration of transplastomic PTD-GFP tobacco and lettuce plants.

<p>Schematic representation of lettuce (A), tobacco (B) chloroplast transformation vectors. LsPrrn, <i>Lactuca sativa</i> rRNA operon promoter and GGAG ribosome binding site; aadA, aminoglycoside 3′-adenylyltransferase gene; smGFP, soluble modified green fluorescent protein; LsTrbcL, 3′ untranslated region (UTR) of <i>Lactuca sativa</i> rbcL gene; LsPpsbA-PTD, promoter and 5′ UTR of <i>Lactuca sativa</i> psbA gene fused to protein transduction domain (PTD, amino acid sequence – RHIKIWFQNRRMKWKK) of pancreatic and duodenal homeobox factor-1 (PDX-1); LsTpsbA, 3′ UTR of <i>Lactuca sativa</i> psbA gene; NtPrrn, <i>Nicotiana tabacum</i> rRNA operon promoter and GGAG ribosome binding site; NtTpsbA, 3′ UTR of <i>Nicotiana tabacum</i> psbA gene; 16S trnI in pLs PTD-GFP, the homologous long flanking sequence from <i>Lactuca sativa</i> chloroplast genome containing 16S 3′ end sequences and full length trnI gene; trnA 23S in pLs PTD-GFP, the homologous long flanking sequence from <i>Lactuca sativa</i> chloroplast genome containing full length trnA gene and 5′ end of the 23S ribosomal RNA subunit; 16S trnI in pLD PTD-GFP, the homologous flanking sequence from <i>Nicotiana tabacum</i> chloroplast genome containing 16S 3′ end sequences and full length trnI gene; trnA in pLD PTD-GFP, the homologous flanking sequence from <i>Nicotiana tabacum</i> chloroplast genome containing full length trnA gene. (C and D) Southern blots of PTD-GFP lettuce and tobacco plants. UT, Untransformed; 1 and 2, transplastomic lines. (E and F) GFP fluorescence in transplastomic PTD-GFP tobacco and lettuce leaves observed under blue light or chlorophyll fluorescence in untransformed control. (G) Non-denaturing gel for quantification of PTD-GFP. GFP standard protein was loaded at indicated concentration (ng). Total soluble proteins from transplastomic plants were extracted three times from independent lines, loaded as indicated (μg) and quantified using densitometry.