title: The development of a high-yield recombinant protein bioreactor through RNAi induced knockdown of ATP/ADP transporter in Solanum tuberosum.
abstract: There is an increased need for high-yield protein production platforms to meet growing demand. Tuber-based production in Solanum tuberosum offers several advantages, including high biomass yield, although protein concentration is typically low. In this work, we investigated the question whether minor interruption of starch biosynthesis can have a positive effect on tuber protein content and/or tuber biomass, as previous work suggested that partial obstruction of starch synthesis had variable effects on tuber yield. To this end, we used a RNAi approach to knock down ATP/ADP transporter and obtained a large number of transgenic lines for screening of lines with improved tuber protein content and/or tuber biomass. The initial screening was based on tuber biomass because of its relative simplicity. We identified a line, riAATP1-10, with minor (less than 15%) reduction in starch, that had a nearly 30% increase in biomass compared to wild-type, producing both more and larger tubers with altered morphological features compared to wild-type. riAATP1-10 tubers have a higher concentration of soluble protein compared to wild-type tubers, with nearly 50% more soluble protein. We assessed the suitability of this line as a new bioreactor by expressing a human scFv, reaching over 0.5% of total soluble protein, a 2-fold increase over the highest accumulating line in a wild-type background. Together with increased biomass and increased levels in total protein content, foreign protein expression in riAATP1-10 line would translate into a nearly 4-fold increase in recombinant protein yield per plant. Our results indicate that riAATP1-10 line provides an improved expression system for production of foreign proteins.
authors.affiliation: Department of Biology, University of Western Ontario, London, Ontario, Canada.
authors.forename: Reynald
authors.initials: R
authors.lastname: Tremblay
authors.forename: Hong
authors.initials: H
authors.lastname: Diao
authors.forename: Norm
authors.initials: N
authors.lastname: Hüner
authors.forename: Anthony M
authors.initials: AM
authors.lastname: Jevnikar
authors.forename: Shengwu
authors.initials: S
authors.lastname: Ma
mesh_terms: D001483:Base Sequence; D018533:Biomass; D019149:Bioreactors; D001709:Biotechnology; D004591:Electrophoresis, Polyacrylamide Gel; D055785:Gene Knockdown Techniques; D006801:Humans; D000226:Mitochondrial ADP, ATP Translocases / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D035281:Plant Tubers / Q000737:chemistry / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D057127:Single-Chain Antibodies / Q000235:genetics / Q000378:metabolism; D011198:Solanum tuberosum / Q000235:genetics* / Q000378:metabolism; D013213:Starch / Q000378:metabolism
chemical_list: D010940:Plant Proteins; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; D013213:Starch; D000226:Mitochondrial ADP, ATP Translocases