Department of plant breeding – abiotic stress group Physiology and genetic variation of nitrogen use efficiency in spinach (Spinacia oleracea L.) July 2015 MSc thesis plant breeding Remco Overeem Registration number: 900605-637-100 Wageningen University The Netherlands Physiology and genetic variation of nitrogen use efficiency in spinach (Spinacia oleracea L.) A literature and experimental study to gain insight in the physiology and genetic variation of nitrogen use efficiency in cultivated spinach (Spinacia oleracea L.) Remco Overeem Registration number: 900605-637-100 MSc thesis plant breeding (PBR-80436) Supervisors: Ir. J. Rafael Chan Navarrete Dr. C. Gerard van der Linden Prof. Dr. Ir. Edith T. Lammerts van Bueren Dr. Ir. Oene Dolstra Examinors: Dr. C. Gerard van der Linden Prof. Dr. Ir. Edith T. Lammerts van Bueren Wageningen University Laboratory of plant breeding Abiotic stress group Droevendaalsesteeg 1 6708 PB Wageningen The Netherlands Table of contents Acknowledgement .................................................................................................................................. 5 Abstract ................................................................................................................................................... 6 1. Introduction .................................................................................................................................... 7 1.1. Nitrogen .................................................................................................................................. 7 1.2. Spinach .................................................................................................................................... 8 1.3. Nitrogen use efficiency ........................................................................................................... 8 1.3.1. Nitrogen uptake and utilization efficiency ..................................................................... 9 1.3.2. Breeding for NUE ............................................................................................................ 9 1.4. Hydroponics and Ingestad model ......................................................................................... 10 1.5. Problem statement ............................................................................................................... 10 1.6. Research objectives and questions ...................................................................................... 10 2. Literature study ............................................................................................................................ 12 2.1. The origin of spinach ............................................................................................................ 12 2.2. Nitrogen use efficiency ......................................................................................................... 12 2.2.1. Nitrogen uptake efficiency ........................................................................................... 12 2.2.1.1. External factors ......................................................................................................... 13 2.2.1.2. Internal factors .......................................................................................................... 15 2.2.2. Nitrogen utilization efficiency ...................................................................................... 18 2.2.2.1. Nitrogen reduction and assimilation ........................................................................ 19 2.2.2.2. Nitrogen transport inside the plant ......................................................................... 22 2.2.2.3. Nitrogen accumulation ............................................................................................. 22 2.3. Concluding remarks .............................................................................................................. 25 3. Materials and methods ................................................................................................................ 27 3.1. Pre-screening mapping population ...................................................................................... 27 3.1.1. Plant material and cultural practices ........................................................................... 27 3.1.2. Measurements .............................................................................................................. 28 3.2. Physiology ............................................................................................................................. 29 3.2.1. Plant material and cultural practices ........................................................................... 29 3.2.2. Measurements .............................................................................................................. 29 3.3. Statistical analysis ................................................................................................................. 30 4. Results ........................................................................................................................................... 31 4.1. Pre-screening ........................................................................................................................ 31 4.1.1. General results .............................................................................................................. 31 4.1.2. Genetic variation .......................................................................................................... 34 4.2. Physiology ............................................................................................................................. 37 4.2.1. General results .............................................................................................................. 37 4.2.2. Interaction effects ......................................................................................................... 39 4.2.3. Correlations ................................................................................................................... 42 5. Discussion ..................................................................................................................................... 43 5.1. Considerations ...................................................................................................................... 43 5.2. Segregation mapping population ......................................................................................... 43 5.3. Nitrogen level effect ............................................................................................................. 43 5.4. Effect of nitrogen application method ................................................................................. 45 5.5. Interaction effects ................................................................................................................ 46 5.6. Correlation of traits to NUE .................................................................................................. 47 6. Conclusions ................................................................................................................................... 49 References ............................................................................................................................................ 50 Appendix I. Genotypes pre-screening ......................................................................................... 61 Appendix II. Hoagland solution .................................................................................................... 61 Appendix III. Experimental design pre-screening .......................................................................... 62 Appendix IV. NAR pre-screening .................................................................................................... 63 Appendix V. Experimental design physiology ............................................................................... 64 Appendix VI. NAR physiology Ingestad .......................................................................................... 65 Appendix VII. NAR physiology depletion ........................................................................................ 66 Appendix VIII. Graphs type of plant material effect ........................................................................ 67 Appendix IX. Variation of several traits per cross ......................................................................... 68 Appendix X. Correlation matrix pre-screening ............................................................................. 69 Appendix XI. REML table N level effect .......................................................................................... 70 Appendix XII. REML table application method effect ..................................................................... 71 Appendix XIII. REML table cultivar x N level effect .......................................................................... 72 Appendix XIV. REML table cultivar x application method effect ..................................................... 73 Appendix XV. Additional graphs physiology ................................................................................... 74 Appendix XVI. Correlations harvest 1 ............................................................................................... 86 Appendix XVII. Correlations harvest 2 .............................................................................................