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. 2012 May 16;7(5):e37179. doi: 10.1371/journal.pone.0037179

Figure 1. Diagram of chloroplast transformation vector and Western blots of C. reinhardti transformed with vectors containing codon optimized pfs25 or pfs28.

Figure 1

(A) The codon optimized nucleotide sequences corresponding to EGF domains 1–4 of pfs25 and pfs28 were separately cloned into an open reading frame that is upstream of a TEV protease site and FLAG epitope. Transgenes were integrated at the psbA locus by homologous recombination. Gene expression is driven by the psbA promoter and mRNA is stabilized by the psbA 5′ and 3′ untranslated regions (UTRs). (B) Western blot analysis of purified algae-produced HMGB1 containing a FLAG epitope (lane 1), lysate of untransformed parental strain (lane 2), lysate of C. reinhardtii containing a-pfs25 total protein (lane 3) and soluble protein (lane 4) probed with anti-FLAG mAbs. (C) Western blot using anti-FLAG mAbs of purified algae-produced HMGB1 containing a FLAG epitope (lane 1), lysate of untransformed parental strain (lane 2), lysate of C. reinhardtii containing a-pfs28 total protein (lane 3) and soluble protein (lane 4) probed with anti-FLAG mAbs.