Expression of anti-Tumor Necrosis Factor alpha (TNFa) Single Chain Variable Fragment (scFv) in Spirodela punctata plants transformed with Agrobacterium tumefaciens.

Biotechnology and applied biochemistry

PubMedID: 25786575

Parthasarathy B, P K S, Venkataraman K, Vijayalakshmi MA. Expression of anti-Tumor Necrosis Factor alpha (TNFa) Single Chain Variable Fragment (scFv) in Spirodela punctata plants transformed with Agrobacterium tumefaciens. Biotechnol Appl Biochem. 2015;.
Therapeutic antibodies against TNFa have been considered effective for some of the autoimmune diseases such as rheumatoid arthritis, Crohn's diseases, etc. But associated limitations of the current therapeutics in terms of cost, availability and immunogenicity have necessitated the need for alternative candidates. Single chain variable fragment (scFv) can negate the limitations tagged with the anti-TNFa therapeutics to a greater extent. In the present study, Spirodela punctata plants were transformed with anti-TNFa through in planta transformation using A. tumefaciens strain, EHA105. Instead of cefotaxime, garlic extract (1mg/ml) was used to remove the agrobacterial cells after co-cultivation. To the best of our knowledge this report shows for the first time the application of plant extracts in transgenic plant development. 95% of the plants survived screening under hygromycin. ScFv cDNA integration in the plant genomic DNA was confirmed at the molecular level by PCR. The transgenic protein expression was followed up to 10 months. Expression of scFv was confirmed by immuno-dot blot. Protein expression levels of up to 6. 3% of total soluble protein (TSP) were observed. GUS and GFP expression was also detected in the antibiotic resistant plants. The paper shows the generation of transgenic Spirodela punctuata plants through in planta transformation. This article is protected by copyright. All rights reserved.