Analysis Of Gene Expression For The Gene Families Involved In Sugar Metabolism And Latex Physiological And Biochemistry Parameters In Latex Regeneration

In rubber tree (Hevea brasiliens is), sucrose is transported, with the involvement of sucrose transporters, through a phloem-mediated pathway from photo synthetic leaves into the rubber-producing laticifers. In the latex, sucrose is broken down into glucose and fructose by invertase and sucrose synthase, and then through the glycolytic pathway, the pentose phosphate pathway and the tricarboxylic acid cycle, provides the material and energy for rubber biosynthesis, especially providing the Acetyl coenzyme A, NADPH and ATP. In this study, seven gene family involved in sugar metabolism were investigated for their expression kinetics during the process of latex regeneration. The gene families include the three gene families (hexokinase, glucose-6-phosphate dehydrogenase, citrate synthase) cloned and characterized in this study, and the families we reported earlier: invertase, sucrose transporter, sucrose synthase and phosphofructokinase. In addition, the latex physiological parameters, along with the activity of some key sugar-metabolizing enzymes were analyzed during latex regeneration. The main results are as follows:1. Based on EST database of Hevea brasiliensis, using reference proteins from Arabidopsis thaliana, Ricinus communis and Populus alba as query sequences, we isolated and cloned four HbHK genes (HbHK1-4), four HbG6PDH genes (MG6PDH1-4); and five HbCS genes (HbCS1-5).2. We analyzed the gene expression of all members of the three gene families in six Hevea tissues (latex, leaf, seed, female flower, male flower and bark) using qPCR. Each tissue has one high expression family member:HbG6PDH3and HbG6PDH4are the highest expression genes in latex; HbG6PDHl, HbHKl, HbHK4and HbCS2are the highest expression genes in leaf3. Four softwares (Delta Ct method, Bestkeeper, NormFinder and GeNorm)exported similar results that identify UBC4, ADF, UBC2a, eIF2and ADF4as the top five suitable references, and18S as the least suitable one. The application of the screened references would improve accuracy and reliability of gene expression analysis in latex regeneration experiments.4. After tapping of the two rubber tree varieties (Reken628and RRIM600), all family genes were analyzed for their expression kinetics in the latex samples collected from the rubber trees retapped at different hours. The results showed that the expression of most genes were up-regulated during the process of latex regeneration. 5. Generally, the expression of HbSUT, HbSUS, HbHK, HbPFK, MG6PDH and HbCS genes in Reken628was higher than that in RRIM600.6. The assay of physiological parameters showed that Reken628was less productive than RR1M600in latex yield, but its tree circumference growth is more than3.8cm per year compaired to the RRIM600. The activity of key enzymes in each pathway of sugar metabolism was also higher in Reken628than in RRIM600. In addition, after tapping, the physiological parameters and gene expression were recovered to normal levels in shorter time in Reken628.This paper systematically analyzed the expression patterns of key gene families involved in sugar metabolism in the process of latex regeneration, along with the change of latex physiological parameters and key enzyme activity. The result provided the basis for further revealing the molecular regulation mechanisms of latex regeneratioa...