Light-Dependent Betanin Production by Transformed Cells of Sugar Beet

The objective of this work is to transform sugar beet (Beta vulgaris L. var. altissima) cells using Agrobacterium tumefaciens, a wild octopine strain B6S3, and to study metabolic changes associated with the transformation. From the primary tumours on leaf fragments, two tumour lines were established: one pale green and the other red-violet in colour. The red-violet pigment was identified by means of high-performance liquid chromatography as betanin. Betanin production is strongly light-dependent. To enhance the betanin yield, nutrient media with different carbohydrates like sucrose, a combination of glucose and fructose, or only glucose or only fructose were tested. The selection of carbohydrate affected betanin production and yield. It was observed that the production of betanin per g of dry mass was 20–40 % higher on simple carbohydrates, especially fructose, than on sucrose or the combination of glucose and fructose. However, due to higher biomass production on the medium with sucrose, the highest total yield of betanin was obtained in the presence of sucrose. The most suitable time for possible betanin extraction was between days 7 and 14 for tissue grown on sucrose. The tumour line seems to be promising as an alternative source of betanin as well as a model to study sugar-regulated genes involved in tissue morphology control.
Svrha je rada bila transformirati stanice šećerne repe (Beta vulgaris L. var. altissima) bakterijom divljeg soja B6S3 Agrobacterium tumefaciens, te proučavati metaboličke promjene nakon transformacije. Od primarnih tumora koji su se razvili na zaraženim listovima, uspostavljene su dvije tumorske linije: svijetlozelena i crveno-ljubičasta. Pigment iz crvene linije identificiran je visokotlačnom tekućinskom kromatografijom kao betanin. Proizvodnja betanina ovisila je o svjetlosti. Da bi se pojačao prinos betanina, ispitana je hranidbena podloga s različitim šećerima, i to: saharozom, glukozom i fruktozom te samo s glukozom ili samo s fruktozom. Proizvodnja i prinos betanina ovisili su o dodanom šećeru. Proizvodnja betanina po gramu suhe tvari bila je 20-40 % viša na monosaharidima, posebice fruktozi, nego na saharozi ili kombinaciji glukoze i fruktoze. Međutim, zbog obilnije biomase, najveći prinos betanina ostvaren je na podlozi sa saharozom. Najpovoljnije vrijeme za iskorištavanje betanina je između prvog i drugog tjedna supkulture na saharozi. Zaključno, dobivena tumorska linija prikladan je alternativni izvor pigmenta betanina, a i model za istraživanje gena koji uz pomoć šećera regulira morfologiju biljnih tkiva.