Abstract
Proteinase inhibitors are a group of defense related proteins, natural antagonists of proteinases, induced upon herbivory, play a defensive role against polyphagous insects and phytopathogens. Serine proteinase inhibitor isolated from Cocculus hirsutus (L.) Diels, Cocculus hirsutus trypsin inhibtor (ChTI) was found effective as antifungal, bactericidal and nematicidal agent. Tomato plants expressing ChTI have been developed by agro-infection with almost 27% transformation efficiency. Stable integration and expression of ChTI has been established by polymerase chain reaction (PCR), inhibitory assay and western blot assay. Transgenic plants showed increased fruit yield, antioxidants, phenolics, flavonoids and titratable acidity. Protein extracts of tomato plants inhibited Helicoverpa armigera (gut proteinases up to 40%. Transgenic plants MT2 and JT2 challenged with 2nd and 4thinstar H. armigera (Hubner) larvae, showed delayed larval growth with 100% mortality. The results put together suggest that ChTI is a potential candidate for developing transgenic plant with multiple biotic stress tolerance. Key words: ChTI, Helicoverpa armigera, insect bioassay, transgenic tomato, trypsin inhibitory assay.
Highlights
Environmental stress is one of the major challenges for plants’ growth and productivity
Since ChTI exhibited bactericidal activity when E. coli harbouring pChTI was grown for long periods, the plasmid was isolated from 3h grown culture. pChTI amplified with M13/ChTI specific forward and reverse primers, yielded 0.7 kb and 0.45 kb amplicons respectively (Supplementary Appendix Figure S1-A)
Agrobacterium mediated transformation of ChTI resulted in 25.7% efficiency and positive T0 plants transferred to soil medium, grown under greenhouse conditions
Summary
Environmental stress is one of the major challenges for plants’ growth and productivity. To overcome stress spawned by herbivory, plants up-regulate defense genes encoding for proteins, secondary metabolites, toxic chemicals and repellents (Jamal et al, 2013). Proteinase inhibitors are major defendants, form inhibitory complexes with specific proteinases by irreversible trapping or tight binding interactions (Clemente et al, 2019). Proteinase inhibitor genes are expressed to regulate proteinase activity within the cell (Rustgi et al, 2017), as response to insect damage, mechanical wounding
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