Please use this identifier to cite or link to this item: http://hdl.handle.net/123456789/28287
Full metadata record
DC FieldValueLanguage
dc.contributor.authorHira Kanwal-
dc.date.accessioned2024-04-15T08:44:51Z-
dc.date.available2024-04-15T08:44:51Z-
dc.date.issued2023-
dc.identifier.urihttp://hdl.handle.net/123456789/28287-
dc.description.abstractVibriosis is a bacterial disease that is caused by fish pathogen Vibrio anguillarum. This deadly hemorrhagic disease affects numerous fish species that are economically important for aquaculture, causing high rates of morbidity and mortality. To prevent fish, bivalves and crustaceans from vibriosis, development of cost-effective vaccine is required. Outer membrane protein K (pOmpK) is an immunogenic protein of Vibrio anguillarum and serves as potential candidate for subunit vaccine development. Plants can be used as affordable bio-factories for expression of vaccine antigens. The present research work aimed to optimize the tissue culture conditions for Spinacia oleracea leaf explants and to develop an efficient transformation protocol for Agrobacterium-mediated stable transformation of Spinacia oleracea with OmpK antigen. Sterilization of spinach seeds with 0.2% mercuric chloride provided good results. Full MS media gave maximum regeneration efficiency for spinach plant. For tissue culture, full MS media supplemented with 1 mg/L BAP and 0.5 mg/L IAA showed the highest callus formation efficiency. Optimization of hygromycin antibiotic was performed for Spinacia oleracea; with 20 mg/L being the optimal hygromycin concentration for selection of transformed leaf explants. Successful stable transformation of spinach was carried out with OmpK antigen. Transformed explants were grown on selection media containing suitable concentration of hygromycin. Explants with 3 days co-cultivation time showed 52.94% callus formation efficiency. Transformation was confirmed through PCR using gene specific primers. Transgene expression in plants was analyzed by quantitative real-time PCR (qRT-PCR) in comparison with β-actin gene as control. Protein expression was confirmed through Dot Blot, Western blotting and ELISA. Taken together, successful expression of Vibrio anguillarum OmpK antigen may facilitate the development of edible and cost-effective subunit vaccine against vibriosis. Keywords: Spinacia oleracea, OmpK antigen, Vibriosis, Agrobacterium-mediated stable transformation, PCR, qRT-PCR, Western blotting, Dot blot.en_US
dc.language.isoenen_US
dc.publisherQuaid I Azam university Islamabaden_US
dc.subjectBiochemistryen_US
dc.titleAntibiotic Optimization And Stable Transformation of Spinach With Outer Membrane Protein K Antigenen_US
dc.typeThesisen_US
Appears in Collections:M.Phil

Files in This Item:
File Description SizeFormat 
BIO 7276.pdfBIO 72762.05 MBAdobe PDFView/Open


Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.