Application of a robust microplate assay to determine induced β-1,3-glucanase and chitinase activity in the cotton plant

Resistance is induced in cotton plants as the result of either viral infection or exogenous application of elicitors. Induced resistance can be evaluated by determining the production of β-1,3-glucanase and chitinase in plants as a biochemical parameter. The assays being used for the determination of chitinase and β-1,3-glucanase activity are laborious and not cost-effective, as the reducing sugars produced by the substrates colloidal chitin and laminarin are very expensive. The concentration of both substrates was standardized and reduced to 0.25% from 4% in a modified microplate assay, which appeared to be more effective. The amount of β-1,3-glucanase and chitinase produced was significant and determined by the new modified assay. The sensitivity of the microplate assay was significantly raised approximately one- to twofold. The existing procedure for determining resistance induced in cotton plants by viruses and elicitors is time-consuming and expensive. However, this resistance can be determined by β-1,3-glucanase and chitinase activity through a modified, cost-effective and robust technique. Here the authors present a 96-well polystyrene microplate assay, instead of a quartz cuvette assay, in which a decreased amount of costly substrates was used and a large number of samples were processed in a short time.