Fractional factorial designs: An underutilized tool for rapid in vitro system development

Authors: Niedz, Randall

Submitted to: Methods in Molecular Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/1/2020
Publication Date: 7/17/2021
Citation: Niedz, R.P. 2021. Fractional factorial designs: An underutilized tool for rapid in vitro system development. Methods in Molecular Biology. 2289:23-45. https://doi.org/10.1007/978-1-0716-1331-3_2.
DOI: https://doi.org/10.1007/978-1-0716-1331-3_2

Interpretive Summary: Plant tissue culture is growing plants or parts of plants in sterile containers. A variety of applications are possible depending on the type of plant part and culture method used. For example, the most widely used application is micropropagation, which is the culture and propagation of plant shoots. Micropropagation is used when very large numbers of plants need to be produced. Plant tissue culture systems are complex because of the many factors involved in successfully growing a plant as it needs to be grown. Some important factors include the type and concentration of mineral nutrients, vitamins, sugars, and plant hormones in the artificial culture medium, as well as the culture environment such as light, temperature, and humidity. The traditional approach to determining the effects of all of these is to vary each factor individually. Varying each factor individually is very inefficient, and cannot determine if any of the factors interact, which they almost certainly do in plant tissue culture. To efficiently identify the factors and determine how they interact requires using the principle and methods of design of experiments (DoE), which was developed about 100 years ago by Sir Ronald Fisher. These methods are underutilized in plant tissue culture research. This study explains the type of DoE experiment required to screen many factors simultaneously and identify those with the greatest effects. This type of DoE experiment is called a fractional factorial. A detailed example is provided that includes all of the calculations so that no statistical software is required.

Technical Abstract: Anther and microspore culture for producing haploid plants are very complex systems, and include general effects where the specific effects must be identified and optimized to develop culture systems capable of producing the large numbers of haploids required by breeding programs. These general effects include genotype, physiological state of the source plant, age of the anthers and microspores, pre-culture treatments, culture conditions, and culture media. Design of experiments (DoE) is an experimental approach specifically designed to identify and optimize the multiple factors that make up complex systems, and is ideally suited for developing in vitro systems to produce haploids. The basic DoE strategy starts by screening multiple factors thought to affect the responses being measured. Screening identifies factors with large and small effects. Factors with large effects are used to manipulate the system, and are moved to the DoE optimization phase such as response surface methodology. Factors with small or trivial effects are eliminated from further consideration, and this simplifies the system. The basic concepts of fractional factorial designs and how to use them are explained. Fractional factorials are the most important DoE screening tool and are the first experiments run before DoE optimization experiments. To illustrate the unique properties of fractional factorials, a detailed example is provided that includes all of the calculations so that no statistical software is required.