Plant morphogenesis and genetic transformation of horticultural brassicas

Sparrow, Penelope Amelia Claire (2003). Plant morphogenesis and genetic transformation of horticultural brassicas. PhD thesis The Open University.



This study set out to elucidate the genetic control of in vitro shoot morphogenesis and the production of transgenic plants in horticultural brassicas. Desirable characteristics associated with successful transformation were identified. These include susceptibility to A. tumefaciens, high shoot regeneration potential and the production of multiple shoots in association with a distinct swelling or callus phase. Diallel analysis showed that both high and multiple shoot regeneration are heritable traits, with additive gene effects accounting for 71 % and 77 % of the genetic variation respectively. High susceptibility to A. tumefaciens was shown to have a positive effect on transformation efficiency. A highly significant putative QTL associated with susceptibility to A. tumefaciens was located on linkage group 09 of B. oleracea, and substitution lines confirmed the presence of this QTL. Additive gene effects also accounted for 79 % of the genetic control of susceptibility to A. tumefaciens. The ability to increase both in vitro shoot regeneration potential and susceptibility to A. tumefaciens in subsequent generations, through conventional breeding methods, was successfully demonstrated. Tissue culture blackening was shown to be a critical factor limiting the production of transgenic shoots.

A Brassica oleracea genotype, DH 1012, was identified that produced transgenic shoots at a rate higher than anything previously reported, for the transformation of cotyledonary petioles using A. tumefaciens. The speed and efficiency of regeneration enabled the isolation of transgenic shoots just 4 weeks after inoculation with A. tumefaciens. The efficacy of DH 1012 as a potential model genotype for transformation is demonstrated. A transformation rate of up to 31 % combined with the availability of an associated genetic linkage map will make this genotype very desirable as a potential research and development tool for studying gene function.

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