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. 2018 Jan 9;8:2644. doi: 10.3389/fmicb.2017.02644

FIGURE 1.

FIGURE 1

Expression and localization of DENV E1E2bv. (A) Real time analysis of DENV E1 and E2 mRNA expression in Pichia pastoris transformed with pDENV-E1E2bv. Total RNA was isolated from the methanol-induced bivalent clone of P. pastoris, transcribed to cDNA using reverse primers specific to either E1 (red) or E2 (green) and subjected to qPCR. Total RNA from P. pastoris transformed with the empty expression vector pAO815 was subjected to RT-qPCR in parallel as a negative control (blue). (B) RT-qPCR amplification profiles obtained using total RNA extracted from un-transformed P. pastoris host (blue) or monovalent P. pastoris clones expressing E1 (red) or E2 (green). In (A,B), the italicized Arabic numerals indicate Ct values. (C) Localization of E1E2bv expression. The bivalent P. pastoris clone (orange bars) lysed either before (U) or after (I) methanol induction and separated into the supernatant (S) and pellet (P) fractions was analyzed by His-Sorb ELISA using a dengue specific mAb 24A12. (D) Similar experiment, as in ‘C,’ performed in parallel with monovalent P. pastoris clones expressing E1 (red bars) or E2 (green bars) proteins.