Cloning and expression analysis of 14-3-3 gene from Saccharum spp.hybrids

【Objective】Cloning the 14-3-3 gene of sugarcane,analyzing and predicting its coding protein structure were condueted to provide references for gene function and metabolic regulatory mechanism studies of sugarcane.【Method】In the present study,a cDNA encoding 14-3-3 protein from Saccharum spp.hybrids was cloned and sequenced by RT-PCR technique based on comparing EST sequences of 14-3-3 protein found in sugarcane.The secondary structure,advanced structure and functional activity sites of 14-3-3 protein were predicted by bioinformatics.【Result】The cDNA length was 784 bp,28.88 kDa of molecular weight and 4.79 of isoelectric point.It contained 771 bp of open reading frame encoding a polypeptide of 256 amino acids.Cluster analysis revealed that the sugarcane polypeptide shared more than 90% identity with rice,broomcorn and maize.The binding motif for the target protein consisted of a basic pocket composed of Lys-50,Arg-57 and Arg-131,as well as Tyr-132,within the third and fifth helices.The activity sites for the phosphoserine were also shown in the 60th,65th,188thand 218thserines.Sugarcane 14-3-3 gene transcripts were detected by real time-PCR in different tissues.The results revealed that 14-3-3 gene could express in different tissues of sugarcane.It expressed highly in stalk and meristem,which may be related with glucose metabolism and cell division.In roots,sugarcane 14-3-3 maybe participate into absorption and metabolic of mineral elements.【Conclusion】Sugarcane 14-3-3 gene could act as one of the candidate genes for signal transduction regulating protein.