Agrobacterium-mediated transformation of chitinase gene from the actinorhizal tree Casuarina equisetifolia in Nicotiana tabacum

• Published in: Biológia Vol. 70; no. 7; pp. 905 - 914
• Main Authors: Veluthakkal, Radha, Modhumita Ghosh Dasgupta
• Format: Journal Article
• Language: English
• Published: Cham De Gruyter Open 01.07.2015; Springer International Publishing; Springer Nature B.V
• Subjects: bioassays; Casuarina equisetifolia; Cell Biology; chitinase; disease resistance; functional validation; fungi; Fusarium oxysporum; gene expression; genetic transformation; hyphae; leaves; Life Sciences; Microbiology; Nicotiana tabacum; pathogenesis-related protein; Plant Sciences; plantlets; promoter regions; quantitative polymerase chain reaction; recombinant proteins; reverse transcriptase polymerase chain reaction; Thanatephorus cucumeris; tobacco; transformation; transgenes; trees; ubiquitin; wild plants; Zoology; chitinase; tobacco; functional validation; transformation; pathogenesis-related protein
• ISSN: 1336-9563; 0006-3088; 1336-9563
• DOI: 10.1515/biolog-2015-0114

Genetic transformation of plants offers the possibility of testing hypotheses on the function of individual genes and enables exploration of transgenes for targeted trait improvement. Cloning of the full-length class I chitinase from the Casuarina equisetifolia (CeChi1) was earlier reported by our team. In the present study, tobacco was used as a model system to functionally evaluate the potential of CeChi1 driven by ubiquitin promoter. The pUH-CeChi1 construct was introduced into tobacco by Agrobacterium-mediated transformation and the putative transformants were confirmed for stable gene integration, transgene expression and recombinant protein production using PCR, RT-qPCR, antifungal assays and in planta analysis. The in vitro antifungal bioassay using the total proteins from leaves of transformed plantlets revealed the characteristic lysis of hyphal tips of pathogenic fungi including Trichosporium vesiculosum, Fusarium oxysporum and Rhizoctonia solani. The in planta bioassay of transformed tobacco showed reduced disease symptoms when compared to untransformed wild plants. The study revealed that the class I chitinase isolated from C. equisetifolia can act as a potential gene resource in future transformation programs for incorporating disease tolerance.