SEED PRODUCTION TECHNOLOGY FOR FENUGREEK (Trigonellafoenum-graecum L.) IN THE CANADIAN PRAIRIES SAIKAT KUMAR BASU M. Sc. Botany University of Calcutta (India), 1998 A Thesis Submitted to the School of Graduate Studies of the University of Lethbridge in Partial Fulfillment of the Requirements for the Degree MASTER OF SCIENCE Department of Biological Sciences University of Lethbridge LETHBRIDGE, ALBERTA, CANADA March, 2006 © Saikat K. Basu, 2006 DEDICATION This work is dedicated to the memories of my beloved grandmother Ms. Radharani Basu, who passed away in India while I was doing this research work in Lethbridge, AB Canada. Until her death in February, 2005 she was a constant source of love, inspiration and encouragement for me. Although she did not have formal education she always took great interest in my research work and showered me with her blessings whenever I needed them most. iii ABSTRACT Fenugreek (Trigonella foenum-graecum L.) is an annual legume mainly used as a spice crop in many parts of the world. "Tristar" is a new forage cultivar that requires -120 days to produce mature seed in western Canada where only -100 frost-free days are available. The goal for this study was to reduce maturity duration for the crop through a series of studies on the genetics and agronomic aspects of fenugreek. This two year study suggests that: 1) mutation breeding using Tristar seed as a base population could be successfull; 2) multi-location trials using world accessions exhibited genotype X environment interaction; 3) swathing of plants before freezing temperatures set in; 4) application of phosphate fertilizer increased seed yield and; 5) foliar sprays of chemicals can be used for production of high quality seed. In this study some short duration, high yielding and determinate lines of fenugreek were produced improving the potential for use of fenugreek and the economics of beef production in western Canada. iv ACKNOWLEDGEMENTS I would like to thank my supervisors; Dr. James E. Thomas, Associate Professor, Department of Biological Sciences, University of Lethbridge and Dr. Surya N. Acharya, Senior Research Scientist, Agriculture and Agri-Food Canada Research Centre, Lethbridge, Alberta for their constant support, guidance and encouragement. I would have never been able to complete my thesis without their help and cooperation. I am very grateful for the research support from the Alberta Agriculture Research Institute and the teaching assistantship program at the University of Lethbridge. I also sincerely express my gratitude for the financial assistance provided by the School of Graduate Studies, University of Lethbridge. This assistance was critical for me to pursue my Graduate work at the University of Lethbridge. I am also thankful for the research facilities provided by the above organizations during the course of this study. I extend my sincere gratitude to my graduate committee members Dr. Rene Barendregt, Dr. Igor Kovalchuk and Dr. Manjula Bandara for their valuable suggestions, cooperation and support throughout my study period. I would like to express special thanks to the following organizations and people for their help and support during my research work: University of Lethbridge: All the Professors in the Department of Biological Sciences including the current chair, Dr. Stewart Rood and the academic assistants (Laurie Pacarynuk, Helena Danyk, Katrina White and Joanne Golden) whom I have had the pleasure of working with during the past two years. I thank Prof. David Townsend and Prof. Pamela Adams of the Faculty of Education for giving me excellent training in teaching and Ms. Shelley Ross, Science Librarian, for helping me in library support. Agriculture and Agri-Food Canada Research Centre: Dr. Hector Carcamo and his research group for their help in the entomological study; Dr. Francois Eudes and his research group for their help in providing the critical laboratory chemicals, Dr. Qin Chen, Dr. Haiyan Li, Dr. Rob Graff and James Pruce for their help in the cytogenetic study; Toby Entz for statistical consultation, Dough Friebel and Doug Messenger for their help and support in the agronomy related work; Scott Erickson for his help with crop pathology and Byron Lee for his help in digital microphotography and Scanning Electron Microscopic study. My thanks go to the entire field crops crew and the greenhouse staff for their constant help during my research work. To my friends and relatives: Special thanks are due to my friends particularly Aaron Puhl, Eric Van Galeen, Raj Dass, Franco Nogarin, Sandy Cahoon and their family members. They made my stay in Lethbridge a highly memorable experience. I would also like to extend my deepest gratitude to Dr. (Mrs.) Nina Acharya for all of her help, support, encouragement and advice, so graciously offered and deeply appreciated. Last but not the least, I would like to thank my parents and family members back in India for their love, support and constant encouragements during the past two years. vi TABLE OF CONTENTS Chapter One: Introduction Chapter Two: Background 2.1. History of the crop 2.2. Origin, distribution and different species 2.3. Botanical perspective of fenugreek 1 2.4. Cytogenetic studies and variability in fenugreek lines 1 2.5. Nutraceutical properties of fenugreek 1 2.5.1. Steroidal sapogenins 1 2.5.2. Galactomanans 1 2.5.3. Isoleucine 1 2.6. Functional food aspects of fenugreek 1 2.7. Agricultural aspects of the crop 2 2.7.1. Cultural requirements for fenugreek 2 2.7.2. Application of soil inoculums, fertilizers, organic manures and sludge...2 2.7.3. Application of plant growth regulators 2 2.7.4. Disease organisms affecting fenugreek 2 2.8. Breeding of fenugreek: 2 2.8.1. Selection 2 2.8.2. Mutation Breeding 2 2.8.3. Hybridization 3 Chapter Three: Genetic Studies on Fenugreek 3 3.1. Introduction 3 3.2. Materials and Methods 3 3.2.1. Greenhouse study 3 3.2.2. Field study 3 3.2.3. Scanning Electron Microscopic (SEM) studies 4 3.2.4. Statistical analysis 4 3.3. Results 43 3.3.1. Greenhouse study 43 3.3.2. Field study 5 vii 3.4. Discussion 64 Chapter Four: Agronomic Studies on Fenugreek 70 4.1. Introduction 70 4.2. Materials and Methods 74 4.2.1. Fenugreek seed material 74 4.2.2. Environments 74 4.2.3. Experimental design 77 4.2.4. Statistical analysis 81 4.3. Results and Discussion 83 4.3.1. Study on growing conditions 83 4.3.2. Study on effect of location on seed and forage production 91 4.3.3. Swath versus combine study 94 4.3.4. Fertilizer trial 96 4.3.5. GA3 and foliar spray study 100 Chapter Five: Cytogenetic Studies on Fenugreek Ill 5.1. Introduction Ill 5.2. Materials and Methods 113 5.2.1. Plant material used 113 5.2.2. Assessment of chromosome number 113 5.2.3. Colchicine treatment 113 5.3. Results and Discussion 116 5.3.1. Comparison of chromosome number in EMS treated lines 116 5.3.2. Identification of tetraploid seedlings derived from colchicine treated Tristar seed 121 5.4. Conclusions 127 Chapter Six: Study on Potential Insect Pests of Fenugreek 128 6.1. Introduction 128 6.2. Materials and Methods 130 6.2.1. Greenhouse study 130 6.2.2. Field study 130 6.3. Results and Discussion 132 viii 6.4. Conclusions 148 Chapter Seven: Thesis Summary 149 References 157 Appendices 171 ix LIST OF TABLES Table 2.2.1. Distribution of fenugreek (Trigonella foenum-graecum L.) by country and continent 9 Table 2.2.2. Common species of the Trigonella genus according to Petropoulos (2002) 9 Table 2.3.1. Morphological characteristics of fenugreek 11 Table 2.5.1. Major chemical constituents of fenugreek seed and leaves 16 Table 2.5.2. Fenugreek and its associated medicinal properties 17 Table 2.6.1. Summary of research progress with respect to the functional food aspect of fenugreek 22 Table 3.3.1.1. Percent seedling survival in Mi, M2 and M3 generations when grown under greenhouse conditions in 2004 and 2005 44 Table 3.3.1.2. Ranges of different morphometric parameters of the Mi plants (single plants per pot) generated in greenhouse measured after 95 days of seeding when the plants were sprayd with desiccant (2004 spring) 48 Table 3.3.1.3. Ranges of different morphometric parameters of the M2 plants(single plants per pot) generated in greenhouse measured after 95 days of seeding when the plants were sprayd with desiccant generated in greenhouse (2004 summer) 49 Table 3.3.1.4. Ranges of different morphometric parameters of the M3 plants(single plants per pot) generated in greenhouse measured after 95 days of seeding when the plants were sprayd with desiccant generated in greenhouse (2005 spring) 50 Table 3.3.2.1. Mean growth ± SE (where appropriate) and yield parameters of Mi (bulk) grown under LRC irrigation (2005) 55 Table 3.3.2.2. Mean growth ± SE (where appropriate) and yield parameters of M2 selected lines grown under LRC irrigation (2005). Lines with a determinate growth habit are highlighted 56 Table 3.3.2.3. Mean growth ± SE (where appropriate) and yield parameters of M3 selected lines grown under LRC irrigation (2005). Lines with a determinate growth habit are highlighted 57 x Table 3.3.2.4. Mean growth and yield parameters of M4 selected lines grown under LRC irrigation (2005). Lines with a determinate growth habit are highlighted 58 Table 4.3.1.1. Mean square (MS), degrees of freedom (df) and probability (Pr) of F value for forage yield, seed yield and 1000 seed weight as determined by a fixed model ANOVA. For this purpose, 45 genotypes were tested in two locations (LRC rain-fed and irrigation) over two years (2004 and 2005) 89 Table 4.3.1.2.