title: Seed specific expression and analysis of recombinant human adenosine deaminase (hADA) in three host plant species.
abstract: The plant seed is a leading platform amongst plant-based storage systems for the production of recombinant proteins. In this study, we compared the activity of human adenosine deaminase (hADA) expressed in transgenic seeds of three different plant species: pea (Pisum sativum L.), Nicotiana benthamiana L. and tarwi (Lupinus mutabilis Sweet). All three species were transformed with the same expression vector containing the hADA gene driven by the seed-specific promoter LegA2 with an apoplast targeting pinII signal peptide. During the study, several independent transgenic lines were generated and screened from each plant species and only lines with a single copy of the gene of interest were used for hADA expression analysis. A stable transgenic canola line expressing the ADA protein, under the control of 35S constitutive promoter was used as both as a positive control and for comparative study with the seed specific promoter. Significant differences were detected in the expression of hADA. The highest activity of the hADA enzyme (Units/g seed) was reported in tarwi (4.26 U/g) followed by pea (3.23 U/g) and Nicotiana benthamiana (1.69 U/g). The expression of mouse ADA in canola was very low in both seed and leaf tissue compared to other host plants, confirming higher activity of seed specific promoter. Altogether, these results suggest that tarwi could be an excellent candidate for the production of valuable recombinant proteins.
authors.affiliation: Prairie Plant Systems, Inc., #1 Technology Road, Box 19A, RR#5, Saskatoon, SK, 7K 3J8, Canada. kmd@prairieplant.com.
authors.forename: Ketan M
authors.initials: KM
authors.lastname: Doshi
authors.affiliation: Prairie Plant Systems, Inc., #1 Technology Road, Box 19A, RR#5, Saskatoon, SK, 7K 3J8, Canada.
authors.forename: Natalia N
authors.initials: NN
authors.lastname: Loukanina
authors.affiliation: National Research Council of Canada-Saskatoon, 110 Gymnasium Place, Saskatoon, SK, S7N 0W9, Canada.
authors.forename: Patricia L
authors.initials: PL
authors.lastname: Polowick
authors.affiliation: Prairie Plant Systems, Inc., #1 Technology Road, Box 19A, RR#5, Saskatoon, SK, 7K 3J8, Canada.
authors.forename: Larry A
authors.initials: LA
authors.lastname: Holbrook
mesh_terms: D000243:Adenosine Deaminase / Q000096:biosynthesis / Q000235:genetics*; D000818:Animals; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D031297:Lupinus / Q000235:genetics; D051379:Mice; D018532:Pisum sativum / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics*; D012639:Seeds / Q000235:genetics*; D014026:Nicotiana / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; C548482:ADA protein, human; D000243:Adenosine Deaminase
keywords: Adenosine deaminase; Lupinus mutabilis; Nicotiana benthamiana; Pisum sativum; Plant; Transformation