title: Ubiquitin promoter-terminator cassette promotes genetically stable expression of the taste-modifying protein miraculin in transgenic lettuce.
abstract: Lettuce is a commercially important leafy vegetable that is cultivated worldwide, and it is also a target crop for plant factories. In this study, lettuce was selected as an alternative platform for recombinant miraculin production because of its fast growth, agronomic value, and wide availability. The taste-modifying protein miraculin is a glycoprotein extracted from the red berries of the West African native shrub Richadella dulcifica. Because of its limited natural availability, many attempts have been made to produce this protein in suitable alternative hosts. We produced transgenic lettuce with miraculin gene driven either by the ubiquitin promoter/terminator cassette from lettuce or a 35S promoter/nos terminator cassette. Miraculin gene expression and miraculin accumulation in both cassettes were compared by quantitative real-time PCR analysis, Western blotting, and enzyme-linked immunosorbent assay. The expression level of the miraculin gene and protein in transgenic lettuce was higher and more genetically stable in the ubiquitin promoter/terminator cassette than in the 35S promoter/nos terminator cassette. These results demonstrated that the ubiquitin promoter/terminator cassette is an efficient platform for the genetically stable expression of the miraculin protein in lettuce and hence this platform is of benefit for recombinant miraculin production on a commercial scale.
authors.affiliation: Graduate School of Life and Environmental Sciences, University of Tsukuba, Tennodai 1-1-1, Tsukuba, Ibaraki, 305-8572, Japan.
authors.forename: Tadayoshi
authors.initials: T
authors.lastname: Hirai
authors.forename: Abdullah Mohammad
authors.initials: AM
authors.lastname: Shohael
authors.forename: You-Wang
authors.initials: YW
authors.lastname: Kim
authors.forename: Megumu
authors.initials: M
authors.lastname: Yano
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Ezura
mesh_terms: D005818:Genetic Engineering; D006023:Glycoproteins / Q000096:biosynthesis* / Q000235:genetics; D018545:Lactuca / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000096:biosynthesis* / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D029632:Sapotaceae / Q000235:genetics; D025801:Ubiquitin / Q000235:genetics*
chemical_list: D006023:Glycoproteins; D010940:Plant Proteins; D011994:Recombinant Proteins; D025801:Ubiquitin; C008533:miraculin protein, Synsepalum dulcificum