Over expression of modified Isomaltulose Synthase Gene II (ImSyGII) under single and double promoters drive unprecedented sugar contents in sugarcane.

Abstract

Sugarcane has been grown all around the world to meet sugar demands for industrial sector. The current sugar recovery percentage in sugarcane cultivars is dismally low which demands scientific efforts for improvements. Multiple approaches were adopted to enhance sugar contents in commercial sugarcane plants in contrast to conventional plant breeding methods. The exploitation of biotechnological methods and exploration of isomaltulose synthetic genes presented a promising solution to increase the existing low level of sugar recovery percentage in Saccharum officinarum L. Isomaltulose synthase gene II was employed and integrated into plant expression vector driven under the leaf and stem specific promoters terminated by nopaline synthase gene in a cloning strategy shown in the present study. Three gene constructs were developed in various combinations driven under promoters Zea mays ubiquitin and Cestrum Yellow Leaf Curl virus in the single and double combined stacked system. The transformation was executed in multiple formats with single transformed events, double promoter transformation events and triple construct stacked promoters in sugarcane induced calli via the particle gene gun. The transformation of ImSyGII in sugarcane genotype HSF-240 was confirmed by molecular gene analysis while expression quantification was determined through Real Time PCR. Furthermore, HPLC was also done to harvest the increased amounts of Isomaltulose in transgenic sugarcane juice. The present work upheld the enhanced ImSyGII expression in leaves owing to the exploitation of ubiquitin, while the Cestrum Yellow Leaf Curl virus promoter enhanced gene expression in sugarcane stems. The employment of three gene constructs collectively produced elite sugar lines producing more than 78% enhancements in whole sugar recovery percentage. The mature internode proved highly efficient and receptive regarding the production of isomaltulose. Quantifications and sugar contents evaluations upheld an increased Brix ratio of transgenic sugarcane lines than control lines.