Expression of Tcchitinase-I gene in transgenic peanut ( Arachis hypogaea L.) confers enhanced resistance against leaf sp
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ORIGINAL PAPER
Expression of Tcchitinase‑I gene in transgenic peanut (Arachis hypogaea L.) confers enhanced resistance against leaf spot and rust diseases Rajinikanth Marka1 · Rama Swamy Nanna1 Received: 25 October 2019 / Accepted: 7 September 2020 © Springer Nature B.V. 2020
Abstract The disease-resistant transgenic peanut cv ICG 13942 plants were developed by using Tcchitinase-I gene. Agrobacterium tumefaciens strain LBA4404 harboring the binary vector (pBinAR) contains the chitinase (Tcchitinase-I) gene and neomycin phosphotransferase resistance (nptII) gene. The transformed shoots were developed on selection medium (MMS + 0.5 mg/L IAA + 15 mg/L TDZ + 100 mg/L Kan + 250 mg/L Cefotaxime) from deembryonated cotyledon (DC) explants. Established plantlets were screened for the presence of Tcchitinase-I and nptII genes. Stable integration and expression of the transgenes (T0) were confirmed by using PCR, RT-PCR and Southern blot analyses. The transformation frequency 63.34% was recorded. All the transformed (T0) plants were found normal, flowered and set seeds. After selfing the T0 plants, a Mendelian inheritance pattern (3:1) for the transgene in T1 progeny is revealed. T 1 transgenic peanut plants were evaluated for resistance against Cercospora arachidicola, C. personatum and Puccinia arachidis by infection with the microspores using detached leaf assay. These T 1 plants have shown longer incubation, latent period and lower infection frequencies in comparison to non-transformed (WT) plants. The Tcchitinase-I gene expression in resistant transgenic plants was compared to that of a susceptible control. A significant negative correlation was recorded between chitinase activity and the frequency of infection to the three tested disease causing agents. Keywords Tcchitinase-I · nptII · DC · Arachis hypogaea · Early leaf spot · Late leaf spot · Rust
Introduction Peanut or groundnut (Arachis hypogaea L.) is an important oilseed crop grown in the tropical and sub-tropical regions of the world. Due to various biotic stresses such as insect pests, bacterial and fungal diseases, the yield in peanut is decreased. Among the major fungal diseases, early leaf spot (ELS) (Cercospora arachidicola), late leaf spot (LLS) (Cercospora personatum) and rust caused by Puccinia arachidis are more destructive (50–70%) in peanut (Subrahmanyam et al. 1984). Chitin is an important cell wall component of fungi and this is degraded by chitinases. Chitinases (E.C. 3.2.1.14) are poly (1,4-(N-acetyl-β-D) * Rama Swamy Nanna [email protected] 1
Plant Biotechnology Research Laboratory, Department of Biotechnology, Kakatiya University, Warangal‑506009, Telangana, India
glucosaminide))-glycanohydrolases. They directly hydrolyze fungal cell wall, chitin the substrate for the enzyme, and by this action fungal hyphallysis and inhibition of fungal growth occur (Patil et al. 2000). Thus, plants use one of the many natural defense mechanisms to resist against pathogens and accumulate proteins (e.g., chitinases) active against disease causi
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