Starch Branching Enzymes and their Genes in Sorghum

Mutisya, J.M. 2004. Starch branching enzymes and their genes in sorghum. Doctor’s dissertation. ISSN 1401-6249, ISBN 91-576-6768-3 Starch is an important raw material both for food and non-food purposes. It is synthesized and stored in source and sink tissues in plants. The starch deposited in amyloplasts of storage tissues possesses several physico-chemical properties, which makes it desirable for diverse applications. For industrial applications, starches high in either amylose or amylopectin are preferred in order to minimize chemical modification. The main purpose in our research is to understand starch synthesis in the sorghum endosperm, and use this information to generate transgenic sorghum with novel starches for both human diet and industrial application. In starch biosynthesis, the activity and expression profile of starch branching enzymes is important in determining both the yield and proportion of amylose and amylopectin. This thesis describes some of the characteristics of starch branching enzymes and their genes in sorghum. The genes encoding two SBE isoforms, SBEIIa and SBEIIb, were cloned and characterised. Comparison of the SBEIIb amino acid sequence with sequences from related species revealed a conserved core stretch of amino acids believed to harbor the catalytic site of the enzyme. Spatial expression patterns showed that the activity of sorghum sbeI and sbeIIb are seed-specific, while sorghum sbeIIa is expressed in endosperm, embryonic and vegetative tissues. We demonstrated that endosperm-specific expression of barley sbeIIb is governed by the second intron of the barley sbeIIb gene, and that the same second intron can confer endosperm specific expression to both sorghum sbeIIb and barley sbeIIa. This implies that a similar regulation factor in barley, which interacts with sequences within the intron, is also present in sorghum. Deletion mutagenesis of the sorghum sbeIIb promoter showed that a, 1-kb fragment 5 ́ of the transcriptional start site constitutes the minimal promoter capable of high level expression. Diurnal expression profiles suggested that an endogenous oscillator in the endosperm controls expression of sbeI, sbIIa and sbeIIb.