| Steroidal saponins as the secondary metaboliteswidely spread in Dioscorea plants,with a pathogens preventation and physiological functions of environmental resistance.In addition,it can be ultilized for synthesizing the steroidal anti-inflammatory drugs,which have a preminent effect in the treatment of cardiovascular and cerebrovascular diseases.Dioscorea Composita,due to its high sapogenin content,high start content,and high yield,is an ideal energy and medicinal non-food plant with high economic and medicinal value.Therefore,it is a great theoretical and practical value to study the steroidal saponins biosynthetic pathway in D.Composita.Based on our transcriptome data of D.camposita and previouse studies about steroidal saponins biosynthesis,this paper has cloned four genes,econding the putative key enzymes of steroidal saponins biosynthetic pathway,CAS,SE,SQS,and HMGR,and make a a preliminary analysis of WRK Y family,which putatively participate in the secondary metabolism regulation of D.camposita.The results obtained are as follows:(1)The key enzyme genes of CAS,SE,SQS,and HMGR have full-length sequences with the complete open reading frames(O RF),which encode the putative proteinswith the highly conserved domains.(2)Full-length sequences of CAS,SE,SQS,and HMG Rwere cloned from D.composita by using the high-fidelity polymerase KD,in addition,the recombinant pCanG-myc vector ofSE or SQS gene were generated.(3)The spatial and temporal expression of CAS,SE,SQS,and HMG Rwere analyzed by RT-PCR,suggesting that,among the four gene,the expression of HMG R in all tissues is the higest,and the expressions of CAS,SE and SQS in different tissues have no significant difference.(4)According the number and combinationof the conserved domains,21 Dc WRK Y were divided into four groups,including Group ?,Group ?,Group?,and undefinded group.(5)Most of the DcWRKY proteins are unstable and basic.Their secondary structure mainly consists of alpha helices,beta folds,extended chains and irregular coils.(6)The subcellular localizations of 21 DcWRKY proteins are mainly in the nucleus,cytoplasm,and mitochondria.(7)The expression of DcWRK Y in different tissues showed that all 21 DcWRKYs have different expression petterns in leaf,stem,and root;the expression of Dc WRK Y6 ?DcWRKY12,and DcWRKY15 were higer than others;the expression of Dc WRKY3 and DcWRKY20 have no significant difference in leaf,stem and root;DcWRK Y5?Dc WRK Y11 and DcWRK Y17 have a highiest expression in stem;the expression of DcWRKY6 and DcWRKY16 in leaf and stem are higher than that in root obviously.This paper take a preliminary analysis for four genes encoded the putative proteins of steroidal saponins biosynthesis and WRK Ys in D.composita The results provide the theoretical basis for further research on steroidal saponin synthetic pathway of D.Composita. |
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