title: Technology of compact MAb and its application for medicinal plant breeding named as missile type molecular breeding.
abstract: Single chain fragment-variable (scFv) enhanced solasodine glycoside accumulation in Solanum khasianum hairy root cultures transformed by the ScFv solamargine (As)-scFv gene. The scFv protein was expressed at a high level in inclusion bodies of E. coli. After being renatured, the scFv protein was purified in a one-step manner by metal chelate affinity chromatography. The yield of refolded and purified scFv was 12.5 mg per 100 ml of cell culture. The characteristics of the As-scFv expressed in E. coli and transgenic hairy roots were similar to those of the parent monoclonal antibody (MAb). The expression of scFv protein provides a low cost and a high yield of functional scFv antibody against solamargine. The full linear range of the ELISA assay using scFv was extended from 1.5-10 µg/ml. The expressed anti-solamargine scFv protein could be useful for determination of total solasodine glycoside content in plant samples by ELISA. Solasodine glycoside levels in the transgenic hairy root were 2.3-fold higher than that in the wild-type hairy root based on the soluble protein level and binding activities. The As-scFv expressed in S. khasianum hairy roots enhanced solasodine glycosides accumulation and provide a novel medicinal plant breeding methodology that can produce a high yield of secondary metabolites.
authors.affiliation: Faculty of Pharmaceutical Sciences, Khon Kaen University, Khon Kaen Thailand, 40002, Thailand. waraporn@kku.ac.th
authors.forename: Waraporn
authors.initials: W
authors.lastname: Putalun
mesh_terms: D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D001947:Breeding; D002846:Chromatography, Affinity; D032621:DNA Shuffling; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004926:Escherichia coli / Q000276:immunology / Q000378:metabolism; D005822:Genetic Vectors / Q000378:metabolism; D006027:Glycosides / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D008517:Phytotherapy; D018517:Plant Roots; D010946:Plants, Medicinal / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D017510:Protein Folding; D057127:Single-Chain Antibodies / Q000032:analysis / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D012991:Solanaceous Alkaloids / Q000032:analysis* / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D032322:Solanum
chemical_list: D000911:Antibodies, Monoclonal; D006027:Glycosides; D057127:Single-Chain Antibodies; D012991:Solanaceous Alkaloids; C026609:beta-solamarine; C012037:solasodine