Associate Professor Richard GardnerAtkinson, Ross G2007-10-172007-10-171993Thesis (PhD--Biology)--University of Auckland, 1993.https://hdl.handle.net/2292/1911Plant biotechnology and molecular biology are now being used to complement conventional breeding programmes in most of the world's important crop species. The aim of the research described in this thesis was to improve New Zealand's horticultural crop plants through application of similar molecular techniques. A) Gene transfer systems for apple, pepino and tamarillo Tissue culture systems were developed for micropropagation and regeneration of apple (cv Royal Gala), pepino (cv El Camino) and tamarillo (selection Oratia Red). In all three species, transient expression of the gusA reporter gene was observed and kanamycin resistant callus was produced, following inoculation with the pKIWI110 binary vector in the avirulent Agrobacterium strain LBA4404. No transgenic apple shoots were obtained. However, transgenic pepino and tamarillo plants expressing the gusA reporter gene, kanamycin resistance and herbicide tolerance were successfully regenerated. PCR and Southern analysis provided molecular evidence for integration of foreign DNA into the genomes of transgenic pepino and tamarillo plants, but indicated deletions of the integrated T-DNAs had occurred with high frequency. Inheritance of the transgenic phenotypes was demonstrated in the progeny of transgenic tamarillo plants. B) Characterisation of polygalacturonase genes A kiwifruit genomic clone with homology to a tomato cDNA clone for polygalacturonase (PG) was sequenced over an 8.1 kb region. The sequence revealed a gene divided into nine exons, with 58% overall identity to the tomato PG gene at the amino acid level. Significant homology was also noted to PG genes isolated from peach, Oenothera organensis and maize, particularly in several blocks of conserved amino acids believed to encode the active site of the enzyme. Analysis of the kiwifruit PG promoter revealed three 81 bp direct repeat sequences just upstream of the kiwifruit peptide start codon. These repeats were also conserved in a second kiwifruit PC genomic clone. Characterisation of partial cDNA clones indicated that at least two mRNAs for PG were expressed in ripe kiwifruit. Southern hybridisation detected the PG gene at low copy number in the genomes of kiwifruit, two other Actinidia species, apple and pepino. PCR was used to amplify a fragment of the apple PG gene for sequence analysis. PG sequences were also used to help define the genetic origin of kiwifruit. A region of the PG gene was amplified and sequenced from four Actinidia species: kiwifruit (A. deliciosa), A. chinensis, A. eriantha and A. chrysantha. These sequences were used to produce a phylogeny using PAUP (phylogenetic analysis using parsimony). Two distinct lineages of PG genes were observed in the genomes of A. deliciosa, A. chinensis and A. eriantha. Within both these lineages, A. deliciosa sequences were quite distinct to those found in the other three Actinidia species, with the exception of a single sequence that was identical to A. chinensis. These results suggest that hexaploid kiwifruit is an allopolyploid with A. chinensis and at least one other Actinidia species as likely progenitors.Scanned from print thesisenItems in ResearchSpace are protected by copyright, with all rights reserved, unless otherwise indicated.https://researchspace.auckland.ac.nz/docs/uoa-docs/rights.htmThesisCopyright: The authorQ112124013