Experimental designs suitable for testing many factors with limited number of explants in tissue culture

Abstract

The majority of plant tissue culture experiments are set up as factorial experiments in completely randomized designs (CRD), randomized complete block designs (RCBD) or split-plot designs (SPD) that require a great number of stabilized cultures. For various reasons, an insufficient number of explants may prevent the employment of CRD, RCBD or SPD with factorial treatments and hinder the optimization of factors affecting culture response. To prioritize the optimization of “the most important” factors affecting culture response, we explored the applicability of Plackett–Burman Design (PBD) and Fractional Factorial Design (FFD) with a limited number of explants. To test the effects of 8 factors (genotype, 6-benzyladenine (BA), CuSO₄ 5H₂O, Fe source, agar, pH, myo-inositol and cold treatment following inoculation of explants) at 2 levels on response of single-node grape vine cultures, 12 treatment combinations were generated according to the PBD and 16 treatment combinations were generated according to the FFD. These designs require many fewer explants since a typical experiment with eight factors, each at two levels of will require 256 (2⁸) treatment combinations. However, the costs of the PBD and FFD are that there is limited, if any information, on interactions and if available may be more difficult to interpret. Of the factors tested, cultivar, BA and agar concentrations were found to be the most important factors affecting culture response. The types of culture response were in agreement with previous reports indicating that PBD and FFD can effectively be employed with limited number of explants to test effects of several factors and prioritize the “most important” factors.