Physiological Response To Exogenous Salicylic Acid On High Temperature Stress And Cloning Of WRKY Gene In Oncidium

Oncidium is a member of Orchidaceae group,originated in the United States,Mexico and other places.Now,Oncidium has become a famous variety with strong competition in international flower market,cultivated in many countries and widely welcomed by consumers.Many varieties of oncidium have bright color flower and also be called dancing girl.The golden color is the most popular variety.With beautiful and lovely shape,Oncidium has a high ornamental value and can be used for home decoration,greening and landscaping in workplace,bouquets processing.High temperature stress is one important problem which need faced in Oncidium production.In this study,we investigated the mechanism of physiological and biochemical responses under high temperature stress by simulating high temperature cultivation environment in artificial climate chamber.After treated by salicylic acid the alleviated action on the heat injury and physiological and biochemical response mechanism of oncidium were studied.One WRKY family transcription factor has been cloned and analysed.It was constructed on the plant expression vector p CAMBIA13 02 by In fusion.The vector was transferred into GV3101.The main conclusions are as follows.1.Oncidium was grown under high temperature cultivation environment at 40 ?(day)/3 0?(night)in artificial climate chamber.Obviously changes were found in leaf color,the upper part of leaf was yellowish,the base showed different degrees of brown.Spraying different concentrations of salicylic acid had a protective effect on the plants under the high temperature stress,and the 1-5mmol/L treatment concentration was suitable.2.Under high temperature stress,40?(day)/30?(night),the activity of superoxide dismutase(SOD)of Oncidium increased,the activity of peroxidase(POD)and catalase(CAT)decreased.The content of soluble sugar,soluble protein and proline increased;the conductivity and malondialdehyde(MDA)content increase continually;the content of glutathione(GSH)decreased.With spraying different concentrations of salicylic acid,the changes of antioxidant enzyme activity were relatively slight,the content of osmotic adjustment substance was significantly increased,the degree of damage of cell membrane was relatively light.It indicated that salicylic acid has a certain alleviated action of high temperature damage on oncidium.3.Under high temperature stress,40?(day)/30?(night),the chlorophyll content of Oncidium decreased.The maximum fluorescence(Fm),the initial fluorescence(Fo)and the maximum photochemical efficiency(Fv/Fm)were all reduced.The values of qP,qN,ETR and(?PS? were all reduced.It was clear that high temperature inhibited the photosynthesis of Oncidium.With spraying different concentrations of salicylic acid,the corresponding fluorescence parameters decreased very slowly in oncidium leaves under the same high temperature treatment.The results indicated that salicylic acid could help maintain the stability of chlorophyll content and photosynthesis in leaves under the high temperature condition,and could dissolve the excess light energy by increasing the qN to avoid the burning of the leaves.4.A WRKY gene,OnWRKY1,was cloned from the leaf of Oncidium using RT-PCR and rapid amplification of cDNA ends.The open reading frame is 1215 bp which is capable of encoding 404 amino acids.Fluorescence quantitative PCR result showed that OnWRKY1 was associated with heat resistance.The sequence was constructed into plant express vector pCAMBIA1302.This recombinant vector pCAMBIA1302 with OnWRKY1 gene was confirmed by PCR and sequencing analysis and the results showed that the recombinant plant vector was constructed successfully.The recombinant plant expression vector was successfully introduced into Agrobacterium tumefaciens GV3101 by freeze-thawing method.