Understanding the breeding systems of Cakile edentula, Cakile maritima (Brassicaceae) and their hybrids Chengjun Li (ORCID: orcid.org/0000-0001-6127-7423) Doctor of Philosophy September 2017 School of Biosciences The University of Melbourne This thesis is being submitted in the total fulfilment of the nominated degree and the research project is sponsored by the China Scholarship Council (CSC) (grant number: 201306240013). Abstract Invasive plant species and their influences have been a frequent topic of discussion in recent decades; however, most studies treat invasive species individually and their arrival and spread as independent events. In reality, species invade sequentially, building in number and interacting. When these species are closely related, they may hybridise. This thesis explores a case study of two such invasive species, Cakile maritima and Cakile edentula in Australia and in particular the roles that the breeding system has played in hybridisation and invasion. The sea rockets, Cakile edentula and C. maritima (Brassicaceae), are closely related and hybrids can sometimes be found between the two. They have contrasting breeding systems: C. maritima is self-incompatible and therefore is an outbreeder, whereas C. edentula is self-compatible. Seeds at different distances from the invasion front of the two species, Cakile maritima and Cakile edentula, were collected and used to produce hybrids, including F1s, F2s and backcrosses. Hybrids and the parental species were also used to assess their relative fitness in a common garden experiment. Results showed that hybrids were successfully produced in both reciprocal directions but differences were observed. Self-incompatibility was inherited in most hybrids but could be enhanced or reduced, inducing a flexible breeding system. Maternal effects were obvious in hybrids. Backcrosses with the self-incompatible species had higher fitness in all hybrids and even higher than both parental species, indicating the existence of heterosis. This thesis has shown that hybridisation between Cakile species with contrasting breeding systems can be achieved in both directions and the interaction is more likely to occur between hybrids and the self-incompatible species after hybridisation due to the dominance of self- incompatibility and heterosis in hybrids. Therefore, unlike traditional mechanisms of plant invasion, the hybridisation with closely related species and the following interaction between hybrids and their parents can provide the newly arriving species with increased mate availability when it spreads into a new environment, easing the Allee effect and thus helping its establishment. Keywords: Cakile edentula, Cakile maritima, breeding system, hybridisation, self-incompatibility, self-compatibility, fitness i Declaration This is to declare that: (i) The thesis comprises only my original work towards the PhD. (ii) Acknowledgement has been made in the text to all other material used. (iii) The thesis is fewer than the maximum word limit in length, exclusive of tables, maps, bibliographies and appendices. ii Acknowledgements First, I would like to acknowledge my supervisor, Professor Roger Cousens, whose patience with me and enthusiasm for my research seems boundless. I am honoured to have had him as my advisor. I also would like to acknowledge my co-supervisors, Dr Peter Ades and Dr Mohsen Mesgaran, for their insightful comments and supports with my research. Meanwhile, I would like to express my great gratitude to all the members of my academic committee, Susan Hoebee, Ed Newbigin and Phillip Salisbury, who were always patient with my questions and provided helpful feedback. I could not make it without their valuable help and insightful suggestions. I would like to thank the nursery and glasshouse technical officers at the University of Melbourne, especially Nicholas Osborne, who provided excellent care of my experiments at Burnley. Your help and kindness made my research enjoyable. In addition, I would like to acknowledge the laboratory technicians and staff at the Burnley Campus who allowed me to use the lab equipment and provided excellent support for all the lab related issues during my study. I thank my fellow lab mates and PhD students, Sara Ohadi and Lynda M. Hanlon, for the stimulating discussion, support and more importantly for all the fun we have had together. I would like to thank all my friends in Melbourne, whose support is precious to my life and research. Last but not the least, I would like to thank my family: my parents and my wife, Qi Sun, for supporting me throughout my PhD and in all aspects of my life. Thank you so much and I love you all! iii Table of Contents GENERAL INTRODUCTION .............................................................................. 1 1.1 INTRODUCTION .................................................................................................................. 1 1.2 HYBRIDISATION AND ITS CONSEQUENCES ............................................................................... 2 1.3 DETERMINANTS OF HYBRIDISATION ....................................................................................... 4 1.4 MODEL PLANT SPECIES ........................................................................................................ 6 1.5 CONCLUSIONS ................................................................................................................... 8 LITERATURE REVIEW ...................................................................................... 9 2.1 FLORAL DEVELOPMENT ....................................................................................................... 9 2.1.1 Early development ................................................................................................. 9 2.1.2 Stigma receptivity and pollen ripeness ................................................................ 11 2.2 PLANT BREEDING SYSTEM .................................................................................................. 13 2.2.1 Pollen post-arrival processes ............................................................................... 13 2.2.2 Self-incompatibility .............................................................................................. 14 2.2.3 Self-compatibility ................................................................................................. 18 2.2.4 The SI × SC rule .................................................................................................... 20 2.2.5 Partial and cryptic self-incompatibility ................................................................ 21 2.2.6 Inheritance of breeding system ........................................................................... 23 2.3 PROGENY FITNESS ............................................................................................................ 25 2.3.1 Seed traits ............................................................................................................ 25 2.3.2 Progeny growth and survival ............................................................................... 27 2.3.3 Reproductive attractiveness and output ............................................................. 29 2.4 CONCLUSIONS AND RESEARCH QUESTIONS ........................................................................... 30 GENERATION OF F1, F2 AND BACKCROSSED HYBRIDS ................................... 33 3.1 INTRODUCTION ................................................................................................................ 33 3.2 PRODUCING F1 HYBRIDS ................................................................................................... 34 3.2.1 Seed collections ................................................................................................... 34 3.2.2 Seed treatments .................................................................................................. 35 3.2.3 Crossing plan for F1s ........................................................................................... 36 3.2.4 Methodology of pollination ................................................................................. 38 3.3 PRODUCING F2 AND BACKCROSSED HYBRIDS ........................................................................ 39 3.3.1 Seed collections ................................................................................................... 40 iv 3.3.2 Seed treatments .................................................................................................. 40 3.3.3 Crossing plan for BCs ........................................................................................... 41 3.3.4 Crossing plan for F2s ........................................................................................... 42 3.4 CONCLUSIONS ................................................................................................................. 43 THE ROLE OF FLORAL DEVELOPMENT IN HYBRIDISATION BETWEEN C. MARITIMA AND C. EDENTULA ............................................................................................ 44 4.1 INTRODUCTION ................................................................................................................ 44 4.2 MATERIALS AND METHODS ................................................................................................ 46 4.2.1 Bud development in Cakile .................................................................................. 46 4.2.2 Anther dehiscence ..............................................................................................