Differential Expression of Isopentenyl Transferase and Cytokinin Oxidase/Dehydrogenase During Pod and Seed Development in Brassica
Thesis DisciplineCellular and Molecular Biology
Degree GrantorUniversity of Canterbury
Degree NameMaster of Science
Consistency of yield and quality of seed are traits not yet optimised by the brassica seed industry in New Zealand. As of 2008, seed producers in Canterbury, New Zealand, exported approximately $18m of brassica seed. However, there is a need to increase both seed quantity and/or quality. The plant hormone group, the cytokinins, regulates many stages of plant growth and development, including cell division and enhancement of sink strength, both of which are important processes in seed development and embryonic growth. The two gene families targeted in this project play a key role in maintaining cytokinin homeostasis. Isopentenyl transferase (IPT) catalyzes the rate limiting step in the formation of cytokinins, and cytokinin oxidase/dehydrogenase (CKX) irreversibly inactivates cytokinins. The aim of this project was to identify those cytokinin gene family members expressing specifically during the early phases of pod and seed development. Initially this study used a rapid-cycling Brassica rapa (RCBr) because of its rapid life cycle, then, as the project developed, a commercial crop of forage brassica (B. napus) was studied. Reverse transcriptase PCR (RT-PCR) and BLAST analysis was used to identify putative IPT and CKX genes from RCBr and B. napus. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was used to measure the expression of individual gene family members during leaf, flower, pod and seed development. BrIPT1, -3, and -5, and BrCKX1, -2, -3, and -5/7 and were shown to express differentially both temporally and spatially within RCBr root, stem, leaf, seed, and pod tissues. BnIPT1, 3 and 7 and BnCKX1, 2, 5 and 7 were also differentially expressed. Particularly strong expression was shown by BrIPT3, BrIPT5 and BrCKX2 in developing seeds. Both Brand BnIPT3 expressed strongly in maturing leaves. In normal plant growth and development, biosynthesis and metabolism of cytokinin is tightly regulated by the plant. Increasing the levels of cytokinins during seed development, either by over expressing IPT3 or IPT5, or decreasing the expression of CKX2, or both, could potentially increase both seed yield and seed vigour.