Use of the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique to analyse the Anthocyanidin Synthase (ANS) gene Locus in Zimbabwean sorghum landraces with different seed proanthocyanidin profiles

Studies on the effects of mutations within flavonoid pathway genes on the resultant flavonoid profiles in sorghum are important in the identification and characterisation of varieties with nutritionally superior flavonoid profiles. In this study, we aimed at determining the effect of mutations at one important flavonoid pathway locus, the anthocyanidin synthase (ANS) gene, on grain flavonoid profile in sorghum. Sequence polymorphisms at this locus were determined in sorghum varieties with different seed proanthocyanidin profiles. The proanthocyanidin profiles of 61 local landraces were determined by the DMACA stain and butanol-HCl assay. The Anthocyanidin synthase (ANS) gene was then amplified using PCR from a subset of 11 landraces, and the amplicons subjected to sequence polymorphism analysis using the restriction fragment length polymorphism (RFLP) technique. Results show that 89% of the brown landraces, 4% of the red and none of the white landraces had detectable proanthocyanidins in their grain. Grain proanthocyanidins ranged from 0.1 to 1.8 AU at 550 nm per gram of sample. Using the PCR-RFLP technique, no sequence variations were detected at the ANS locus. Consequently, the different proanthocyanidin profiles observed could not be attributed, according to the methods used, to events at the ANS gene locus. These could be due to mutations at other loci or a combination of genetic and environmental factors.   Key words: Sorghum, flavonoid, flavonoid profile, mutation, restriction fragment length polymorphism (RFLP), condensed tannins, Zimbabwe.