Preliminary Studies On Hyperaccumulating Plant Phytolacca Responses To Heavy Metal Stress Comparative Proteomics

Heavy metals have a poor mobility, long residence time, and cannotbe degraded by microorganism in the environment. With the development of theindustrial economy,the heavy metal pollutioncan be caused by the mining,manufacturing, sewage, irrigation, the use of heavy metals products or contaminantscontaining heavy metal and other human factors, and it have become major ecologicalproblems. In recent years, the prevention and repair of heavy metal pollution hasbecome a top priority to environmental protection, andphytoremediationbased onhyperaccumulators is increasing concerned. Previous studies have found Phytolaccacan hyperaccumulate cadmium, manganese and zinc, thus it became one of the mostpopular materials in the field of environmental biotechnology. Since some works onhyperaccumulating mechanism have been reported, less research can be foundinproteomics.Therefore, it is necessary to carry out Phytolacca proteomics research.Inthis thesis,Phytolaccawas used as materials in the laboratory hydroponic cultureconditions under heavy metals cadmium or manganese stress, and a system oftwo-dimensional electrophoresis of Phytolaccawas built. The main results of thispaper are as follows:Phytolaccawas stressed by cadmium or manganese.Phytolacca under cadmiumstress has appeared yellow, slow growth, accelerated aging and other serious toxicsymptomsat72hours, and more than100μM high concentrations of Cd ismoreserious,even it lead to the death of plantsin a short time. Phytolacca under5mM ofMn2+stress grew normally, but stress of more than10mM Mn2+can lead leaves turnwhite, malformations, and the higher the concentration of Mn2+, the more serious theplant being poisoned.In this study, optimized operation condition of2-DE mainly included the samplepreparation of Phytolacca stem and leaf tissue and isoelectric focusingprocedures.TCA-acetone method was used to extract protein.In the experiment, we used the optimized system of two-dimensionalelectrophoresis to extractsuccessfully total protein of Phytolacca stems and leavestissue under200μM Cd2+stress at0h,12h,24h,48h,72h,and obtained five group 2-DE gels having good reproducibility.Every group matching rate had reached morethan85%.63significantly different proteins were found by comparing the2-DEgelsof different stress time points.(1) By comparing2-DE gels in0h-24h, we found36differentialspots, of which13spots were up-regulated,23spots were down-regulated.13up-regulated spotscontinued upward in0h-12h-24h, the expression level of some points raised at the12hand24h decline.23protein spots were continuously down-regulated at most timeduring0h-12h-24h, and some points expression levels appear downwards at12handthen increased, and be unchanged or increased at12h, increased to24h and then be indownward trend.(2) After analysing2-DE gelsat0h,24h,48h,72h, a total of59significantlydifferent(p<0.05) pointswas found.28spots were up-regulated,28spots weredown-regulated,3spots were special (disappear or appear). In these points,60%significantly up-regulated spots began to increaseat12h, continued to increase at24h.70%significantly down-regulated spots began to decrease at12h or24h and thencontinued to fall.