Production of microbial mutan polysaccharide by expression of a mutansucrase gene (gtfI) in sugarcane
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Due to its high productivity and sucrose content, sugarcane (Saccharum officinarum) is becoming the source of high-value bioproducts. Expression of bacterial extracellular polysaccharide genes in non-biopolymer accumulating plants is an excellent resource for production of added-value products. To this end, an expression cassette containing a full-length glucosyltransferase (gtfI) gene from Streptococcus downei driven by a CaMV promoter was expressed in a commercial sugarcane cultivar (CP48-103) using a biolistic approach. Copy number was assessed for a number of selected transgenic sugarcane lines by DNA blot analysis, where it was corroborated that each transgenic line contained at least two gtfI copies. The southern blot analysis of gtfI-expressing lines showed that the number of integrated copies ranged from two to four. The expression of gtfI in transgenic sugarcane plants was confirmed by mRNA blot analysis and qRT-PCR analysis. The expression of gtfI in transgenic sugarcane plants resulted in an approximate 30% reduction in sucrose accumulation, suggesting that mutansucrase actively converted sucrose to mutan polymer. In internodal stalk tissues, mutan polymer accumulated up to 55.9 mg/g FW, which apparent through glucan staining. The levels of glucose and fructose increased nearly by twofold, suggesting that mutansucrase may also have hydrolyzing activity.
KeywordsExopolysaccharide Glycosyltransferases Sucrose, Sugarcane Transgenic plants
We are grateful to Professor Richard G. F. Visser from Wageningen University for his critical reading of the manuscript and valuable feedback. We also would like to express appreciation to Ahwaz Sugarcane Research Center staff for their excellent assistance.
Authors contribution statement
M.A designed and performed the experiments. F.N-F designed and performed some of the experiments, and wrote the manuscript. A.A and B.B helped in designing some of the experiments. S.K critically read the manuscript and helped in designing experiments.
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