Transient Expression Analysis of Agave Sisalana Heat Shock Protein Gene (ashsp70) in Model Species (nicotiana Benthamiana) under Heat Stress

This study is aimed at investigating whether overexpression of the heat shock protein gene (AsHSP70) isolated from Agave sisalana will improve the heat stress tolerance in plants. AsHSP70 gene with a full-length cDNA of 1800 bp, encoding 282 amino acids, was functionally characterized. Bioinformatics tools showed the theoretical isoelectric point is pI = 4.780, located in the nucleus with 76.8% reliability having an N-terminal signal domain. 3D model, showed an ATP and ADP ligands with –6.2 and 6.5 kJ/mol binding affinity respectively. AsHSP70 was cloned in Plant expression vector in fusion with Green Florescent Protein from pGWB5 vector. Syringe agroinfiltration of Agrobacterium tumefaciens harboring AsHSP70/GFP was done into leaf of Nicotiana benthamiana. Spermidine was applied to the infiltrated tobacco plants for exogenous induction of polyamines under heat stress. Relative expression of AsHSP70 was significant in tobacco leaf under heat stress. Membrane stability index was about 95.59 and 96.53% and leaf relative water content was 4.43 and 3.44% respectively in infiltrated and polyamine-induced plants. Proline and sugar content were also increased under heat stress. Hence, we established that AsHSP70 is expressing transiently under heat stress in tobacco as model species and further genetic transformation of this gene will lead to induce the stress tolerance in agriculturally important crops.