ASPECTS OF THE REPRODUCTIVE BIOLOGY, BREEDING ~ ' \ SYSTEM AND HORTICULTURAL IMPROVEMENT OF THE GENUSPANDOREA submitted by Elizabeth Ann James B.Sc.(Hons) (LaTrobe) A thesis submitted in total fulfilment of the requirements for the degree of Master of Science School of Botany University of Melbourne Parkville, Melbourne, 3052 Australia April, 1992 Pandorea pandorana (GS) ii DECLARATION Except where reference is made in the text of the thesis, this thesis contains no material published elsewhere or extracted in whole or in part from a thesis presented by me for another degree or diploma. No other person's work has been used without due acknowledgement in the main text of the thesis. This thesis has not been submitted for the award of any other degree or diploma in any other tertiary institution. .. .\.~: ..Lt.92- E. A. James 111 ACKNOWLEDGEMENTS The experiments in this thesis were carried out at the Institute of Plant Sciences (IPS) as part of a research program of the Department of Food and Agriculture (DoPA). I would like to thank the following people for their support and guidance: My supervisor, Prof. R. Bruce Knox, Botany School, and Dr. David Beardsell, Dennis Richards and Bill Thompson (IPS) for their most welcome advice and comments and open door policy. Dr. Graeme Frith, Director of IPS for permission to pursue these studies in the Botany School at the University of Melbourne. Dr. John Reynolds and Peter Franz, Biometricians (DoPA), for statistical advice far beyond the call of duty. Dr. Robert Bertin, College of the Holy Cross, Massachussetts, USA and Dr. Alwyn Gentry, Missouri Botanic Gardens, Missouri, USA for advice and information on the family Bignoniaceae. Philip Taylor for sharing his expertise in electron microscopy. Drs Gordon Guymer and Bill McDonald, National Herbarium, Queensland, Elizabeth Brown, National Herbarium, NSW and Alexander Buchanan, National Herbarium, Tasmania, for their assistance in the location of plant material. Mr. Col Harman, O'Reilly's Guest House, Green Mountain, Qld for leading me to P. baileyana. Peter and Ann Radke (Yuruga Nursery, Tolga), Bruce Gray (CSIRO, Atherton), David Beardsell and Tony Slater for some additional plant material. Dr. Yvonne Fripp and Harald Hozmec, Department of Genetics and Human Variation, LaTrobe University, for advice and technical assistance with the isozyme analyses. Francha Horlock and Michelle Bankier for technical assistance and the maintenance of my Pandorea collection. Jocelyn von Kanel and Dr. Doris Blasing for translations of French and German manuscripts. In addition to those 1nentioned above, I would like to thank my colleagues at IPS and the Botany School for their friendship and encouragement especially , Trish Grant, Roger Ashburner, Vijay Kaul and Jan1es Hutchinson. My family, especially Gavan, Davis and Arlie, thanks tean1! iv REASONS FOR THIS STUDY The ornamental plant industry relies on the introduction of new products with unusual or novel attributes. Conventional plant breeding techniques in conjunction with mutation breeding and the selection of natural mutations have been the major sources of new material for the ornamental plant industry. The Australian tlora has 1nuch potential in the ornarnental plant industry and is being used increasingly by both the cuttlower industry and the nursery industry. The majority of material sold on local and export markets is unirnproved although selection and plantation production of material is increasing. The genus Pandorea has a small local market for two species, P. jasminoides and P. pandorana, in the landscape industry. The former species has large flowers but very little intraspecific variation. The latter species has smaller flowers but considerable variability in both flower size and colour. The two remaining Australian species, P. baileyana and P. nervosa are not in commercial horticulture. The potential to produce elite 1naterial for the horticultural and landscape industries through a breeding program within the genus is good. Interspecific hybrids have been produced in the family between species in the tribe Tecomeae to which the genus Pandorea belongs. This study aims to study aspects of the reproductive biology and breeding system of the genus Pandorea, under experimental conditions, and to develop techniques for the production of interspecific hybrids from selected material. v ABSTRACT The Australian flora represent a potential genetic resource for the production of new cultivars for the local and international ornamental plant industry. The genus Pando rea contains four species in Australia. All are clitnbers with a range of flower colour and size. This study aimed initially to collect a range of genotypes from the Australian species of Pandorea. It then sought to identify and study aspects of the reproductive biology and breeding system of the genus Pandorea. It also aimed to cross species using the basic techniques of plant breeding to provide unique genetic combinations as a basis for new horticultural varieties of Pandorea. Floral dimensions were xneasured and con1pared for all species. The reproductive biology was indicative of a genus comprising obligate outcrossing individuals. Strong self-incompatibility was found in three species. The fourth was not tested. The stigrnas were receptive prior to anthesis. Pollen viability deteriorated over a five day period. Interspecific hybrid seedlings were obtained for some crosses through embryo culture. Embryos aborted if left on the parent plant apparently due to endospenn failure. The success of rescuing interspecific hybrid etnbryos was related to the stage at which embryo develop1nent was arrested. Well-developed cotyledonary einbryos were readily grown in aseptic culture and were acclitnatized with ease fron1 tissue culture. Less mature embryos gern1inated precociously and failed to continue through the normal embryo developrnent. Isozyme analyses were useful for corroborating the hybrid status of so1ne putative hybrid seedlings produced in this study. One hundred and seventeen confirmed interspecific hybrids and seven unconfinned hybrids have been acclimatized to standard nursery conditions. They are anticipated to flower for the first time in the 1994 flowering season when their floral characteristics can be assessed for horticultural potential. vi TABLE OF CONTENTS Page CHAPTER 1: REPRODUCTIVE BIOLOGY, FLORAL STRUCTURE AND PHYLOGENY OF THE BIGNONIACEAE . 1 1.1 SYSTEMATICS AND EVOLUTION .......................... 1 1.1.1 Evolution and phylogeny .............................. 1 1.1.2 Plant and Flower Morphology ........................... 3 1.1.2.1 Habit ...................................... 3 1.1.2.2 Leaves ..................................... 4 1.1.2.3 Indument ................................... 4 1.1.2.4 Inflorescence ................................. 6 1. 1. 2. 5 Calyx . 7 1.1.2.6 Corolla ..................................... 7 1.1.2.7 Stamens .................................... 9 1.1.2.8 Pistil ...................................... 9 1.1.2.9 Disk ..................................... 10 1. 1 . 2. 10 Fruit . 10 1.1. 2.11 Seeds .................................... 10 1.1.1.12 Seedlings . 11 1.1. 3 Cytology of fatni1 y . 11 1.1.4 Chemical constituents ............................... 15 1.2 REPRODUCTIVE BIOLOGY .............................. 16 1.2.1 Pollen ........................................ 16 1.2.1.1 Description of pollen types in the family Bignoniaceae . 16 1.2.1.2 Evolution of pollen type . 19 1.2.2 Pistil ........................................ 20 1.2. 3 Embryology . ............. 21 1.2.4 Conclusions ..................................... 22 vii 1.3 REPRODUCTIVE BIOLOGY IN THE FAMILY BIGNONIACEAE ...... 24 1.3.1 Pollination biology ................................. 24 1.3.2 Influence of pollination on fruit set ....................... 26 1.3.3 Self-incompatibility ................................ 26 1.3.4 Interspecific hybridization ............................ 28 1.3.5 Conclusions ..................................... 29 1.4 AUSTRALIAN BIGNONIACEAE ........................... 29 1.5 GENERAL CONCLUSIONS .............................. 31 CHAPTER 2: MATERIALS AND METHODS ..................... 32 2.1 PLANT MATERIAL USED IN THIS STUDY ................... 32 2.1.1 Reproductive biology and breeding system . 32 2.1.2 Tissue culture .................................... 32 2.2 FLORAL DIMENSIONS ................................. 35 2.3 BREEDING SYSTEM ................................... 36 2.3.1 Reproductive structures .............................. 36 2.3.1.1 Pistil ..................................... 36 2.3.1.2 Pollen .................................... 36 2.3.2 Pollen-ovule ratio ................................. 38 2.3.3 Pollen-tube growth in selfed and outcrossed flowers ............ 38 2.3.4 Separation of male and female function .................... 38 2.3.5 Self-incon1patibility, genotype and reciprocal effects . 39 2.4 INTERSPECIFIC HYBRIDIZATION ......................... 39 2.4.1 Pollinations ..................................... 39 2.4.2 Embryo rescue ................................... 39 2.4.3 Fruit 1neasurements and seed germination ................... 41 2.4.4 Isozy1ne analysis of putative hybrids ...................... 41 2.5 STATISTICAL ANALYSES ............................... 44 CHAPTER 3: THE GENUS PANDOREA ...................... 45 3.1 DESCRIPTION OF THE GENUS ........................... 45 Vlll 3.2 FLORAL MORPHOLOGY ................................ 46 3.2.1 Inflorescence ...................................