Investigation into the functional nature of Frc locus conditioning fructan levels in onion
Thesis DisciplinePlant Biotechnology
Degree GrantorUniversity of Canterbury
Degree NameDoctor of Philosophy
Frc, a major gene on chromosome 8, conditions fructan levels in onions (Allium cepa L). In order to assist genetic dissection of this locus, this study aimed to determine the factors influencing varying fructan levels in high- and low-fructan genotypes. Mapping families were developed and analysed to study the genetic architecture for the fructan trait, and to check the association of the identified variables with the Frc locus. To facilitate reliable and practicable sugar assays in onions, a newly-adapted high-throughput microplate enzymatic assay was validated in this study. The reliability of using leaf sugars as a representative of bulb sugars in a mapping population was studied. Microplate enzymatic sugar assays were carried out on a segregating onion cross to validate the use of maltases in sugar analysis, and the results obtained were validated against HPLC-PAD. Sucrose measured in microplates employing maltases as the hydrolytic enzyme was in agreement with HPLC-PAD results. Maltase enzymes specifically hydrolysed sucrose in onions, providing an alternate tool in place of expensive sugar assay kits. Use of the microplate-enzymatic assay provided a rapid, cheap and practicable method for sugar analysis in onion. Differences in carbohydrate content, sucrose metabolising enzyme activities and their expression levels were monitored in developing leaf blades and leaf bases of four high- and four low-fructan genotypes. The variation in fructan accumulation between high- and low-fructan genotypes was due to the variation in sucrose metabolism. SPS expression and activity did not vary between high- and low-fructan genotypes. Acid invertase and 1-SST showed significant variation in their activities between the two fructan groups. Post-transcriptional and translational regulation of AI and 1-SST respectively, are suggested. Mapping populations analysed for non-structural carbohydrates showed very wide segregation for fructan (80 to 600 g kg⁻¹) and other NSC content, and were well-suited for detailed genetic and physiological analysis. Single marker analysis was carried out to study the association between the combined enzyme activity (CEA; acid invertase + 1-SST) and the Frc markers. Significant association between CEA and Frc markers has suggested genes regulating acid invertases or 1-SST or both underlie Frc. Leaf blade NSC did not correlate with bulb sugars and thus cannot be used as a phenotypic marker for early selection of bulb NSC traits.