Cloning And Function Analysis Of ShHXT1 And ShHXT2 In Sugarcance

Sugarcane(Saccharum officinarum L.)is a high efficiency of C4 plants,which mainly growing in tropical and subtropical areas.It is important sugar crop in the world.Sucrose,the main product of photosynthesis,is the major form of carbohydrate to transport,distribute and store.A part of sucrose is stored,others can not be used directly,need to re-hydrolyzed to monosaccharides,with the assistance of the monosaccharide transporters,which is absorbed and used by sink,to maintain normal physiological function after sucrose is transported from source organs to sink organs.Therefore,the isolation and identification of sugarcane monosaccharide transporters genes will shed light on the distribution mechanisms in sugarcane.1.In this article,the cDNA encoding hexose transporters of sugarcane were cloned by RT-PCR from NCBI.The primers were designed according to the homologous species.The ShHXT1 contains an Open Reading Frame of 1473bp,encoding 460 amino acids and have 10 transmembrane region;ShHXT2 contains an Open Reading Frame of 1623bp,encoding 540 amino acids and have 12 typical transmembrane region.2.The subcellular localization of ShHXT1 and ShHXT2:onion epidermal cells were injected by Agrobacteriurn and the results show that:they belong membrane proteins.3.Yeast mutant EBY.VW4000 was used to identify the frurose uptake activity.4.The expression patterns analysis of ShHXT1 and ShHXT2 in different tissues in sugarcane by fluorescent quantitative PCR and the results show that:ShHXT1 and ShHXT2 are higher expressed in immature tissues in sugarcane,indicating that ShHXTl and ShHXT2 promote growth and development.5.The expression patterns analysis of ShHXT1 and ShHXT2 under cold stress in sugarcane plantlets by fluorescent quantitative PCR and the results show that:the expression levels of ShHXT1 and ShHXT2 gradually increased within 78 hours,indicating the important roles of ShHXT1 and ShHXT2 under cold stress.6.The expression patterns analysis of ShHXT1 and ShHXT2 in three kinds of sugarcane diseased leaves by fluorescent quantitative PCR and the results show that:the expression levels of ShHXTl and ShHXT2 in sugarcane leaves infected by brown streak disease,smut disease and red stripe disease are higher compared with those in healthy sugarcane leaves.These results indicate that they may be activated and provide carbon environment for pathogen invasion and propagation.7.Genetic transformation of tobacco obtained 9 positive ShHXT2 transgenic plants,The expression patterns analysis of ShHXT2 under cold stress show that:indicating the important roles of ShHXT2.8.Genetic transformation of sugarcane through agrobacterium-mediated method obtained 12 positive ShHXT2 transgenic plants.