Use of response surface methodology for optimization of a shoot regeneration protocol in Basilicum polystachyon

Abstract

Response surface methodology (RSM) is a collection of techniques useful for analyzing and optimizing problems where several explanatory covariates influence a response. Although this technique is extensively used in various mixture experiments, its application in standardization of micropropagation protocols is limited. The theoretical developments of RSM are usually concerned with continuous data; hence, linear model theory becomes relevant. In plant tissue culture, in which the response variables are mostly numerical data, the development of RSM in a generalized linear model (GLM) setup is of interest from both a theoretical as well as an application perspective. In the present paper, RSM, as applicable for count data, has been used for modeling, analyzing, and optimizing in vitro regeneration of multiple shoots of Basilicum polystachyon, an important medicinal plant. The specific issues addressed herein are the determination of the optimum concentration of plant growth regulators (i.e., the range of variation in dosages of each covariate) at which the regeneration potential of shoot tip explants is expected to increase, selection of the appropriate growth function (response function) of shoot tip, and determination of the optimum levels of the explanatory variables (i.e., the different combination of dosages of various control factors) for experimentation. According to the present analysis, the optimum level combinations of growth regulators for regeneration of multiple shoots from shoot tip explants of B. polystachyon is 8.19 μM benzyladenine and 2.36 μM naphthalene acetic acid, with a response of approximately 12 regenerated shoots.