Laboratory Exercise Demonstration of Allelopathy of Horseradish Root Extract on Lettuce Seed S Tyler Simpson† Yu-Chun Chiu† Michelle Richards-Babb‡ Jessica M. Blythe§ Kang-Mo Ku †¶* From the †Division of Plant and Soil Sciences, West Virginia University, Morgantown, West Virginia, 26505, ‡C. Eugene Bennett Department of Chemistry, West Virginia University, Morgantown, West Virginia, 26506, §School of Design & Community Development, Davis College, West Virginia University, Morgantown, West Virginia, 26505, ¶Department of Horticulture, College of Agriculture and Life Sciences, Chonnam National University, Gwangju, 61186, Republic of Korea Abstract Allelopathy plays crucial roles in invasive plant viability and agricultural production systems. However, there is no well- established hands-on learning activity to teach the concept of allelopathy. Nor is there an activity which allows students to gain knowledge about glucosinolates and their corresponding enzyme, myrosinase, which are present in almost all Brassica crops. Lettuce germination was counted by the students from three different treatments including water treated with Paraf- ilm sealing, horseradish treated with Parafilm sealing, and horseradish treated without Parafilm sealing. Additionally, let- tuce root length was measured by students using ImageJ software from each treatment using pictures captured by students’ smartphones. Students took an identical quiz as a pre-laboratory and a post-laboratory assignment. Their aver- age scores on the pre-laboratory and post-laboratory quizzes were 3.14 and 6.56 out of 10, respectively, indicating the lab activity significantly improved students’ understanding of allelopathy and glucosinolate-myrosinase system. In addition, students (n = 76) completed a survey post-laboratory to assess their self-efficacy. This simple and cost-effective labo- ratory activity improved students’ knowledge and skill devel- opment as it made learning more inviting, meaningful, and fun. © 2019 International Union of Biochemistry and Molecular Biology, 00(00):1–8, 2019. Keywords: High school/introductory biochemistry; first-year under- graduate/general; laboratory instruction; applications of biochemistry Introduction Plants compete with neighboring plants to secure nutrient resources to grow. Some plants produce and use secondary metabolites that suppress neighboring plants’ growth and development to better position themselves in this competi- tion for resources. This is known as allelopathy, which is defined as the influence (beneficial or detrimental) on a plant or an organism due to the biochemicals (allelochem- icals) produced by a separate plant or organism (either same or another species) [1]. Allelochemicals are exuded from plants by a variety of modes: leaching, root exudation, volatilization, residue decomposition, and other processes. Many invasive species’ distribution and success are depen- dent on allelochemicals, which act as natural herbicides on native species [2]. In addition, the allelopathic mechanism of some cover crops is widely used in current agricultural production systems as a sustainable and organic method to manage weeds and soil fertility. One such example is garlic mustard, which utilizes the allelochemical sinigrin, a gluco- sinolate, to disrupt the germination of other nearby plants [3]. Glucosinolates are plant secondary metabolites that are derived from glucose and an amino acid. After reaction with myrosinase, an endogenous enzyme, glucosinolates are hydrolyzed into glucosinolate hydrolysis products by cleav- ing thio-linked glucose on the structure. Hydrolysis prod- ucts, such as isothiocyanates, are responsible for the pungent flavor of many Brassica vegetables, including for example broccoli, horseradish, mustard, and cabbage. Volume 00, Number 00, Month 2019, Pages 1–8 *To whom correspondence should be addressed. Tel.: (304) 293-2549; Fax: 304-293-3740. E-mail: kangmo.ku@mail.wvu.edu. S Additional Supporting Information may be found in the online version of this article. Received 13 August 2018; Revised 17 December 2018; Accepted 19 January 2019 DOI 10.1002/bmb.21219 Published online 00 Month 2018 in Wiley Online Library (wileyonlinelibrary.com) Biochemistry and Molecular Biology Education 1