Study On New Methods For Rapid Extraction Of Rna From Food-borne Pathogens And MiRNA From Serum

The extraction of ribonucleic acid(RNA)is an important step in the nucleic acid detection process,and the RNA extraction time has a great influence on the efficiency of the nucleic acid detection process.The current RNA extraction methods are time-consuming,complex and easy to cause RNA degradation in the extraction process,which largely limits the efficiency of nucleic acid detection.Therefore,it is urgent to study a new rapid and effective RNA extraction technology to speed up the sample pre-processing time in nucleic acid detection,so as to improve the efficiency of nucleic acid detection.Formamide can protect RNA from degradation and separate RNA from samples;chitosan is positively charged at p H<6.3,and can purify RNA by electrostatic adsorption combined with negatively charged RNA.Therefore,this article for the first proposed a new rapid RNA extraction method,FBS/chitosan,based on formamide separation and chitosan-silica membrane purification.Taking food borne pathogenic bacteria Escherichia coli as the research object,RNA in E.coli was separated by formamide lysate,and then purified by chitosan modified silica membrane.The total RNA could be extracted within15 minutes.Compared with the TRIzol method and commercial kits,the extraction process was simpler,faster,and lower in cost.It had certain development potential and was expected to become a rapid sample preparation tool for nucleic acid detection methods with increasing demand.Based on the hybridization of the immobilized nucleic acid capture probe to the target,the nucleic acid extraction method of specific capture target has been applied.However,the process of nucleic acid passive hybridization generally takes 2 hours or even longer,while the application of a certain electric field strength can accelerate the movement of nucleic acids,thereby accelerating hybridization.Therefore,this article used electric field-assisted accelerated hybridization for the first time to rapidly separate mi RNA,which proposed an ultra-fast mi RNA extraction scheme.By applying an alternating electric field of +0.5V/-0.2V,the hybridization between mi RNA and capture probe was accelerated.The hybrid of the capture probe and the target mi RNA was separated and eluted from the electrode by a negative electric field of-1 V.The whole process only needed 150 s to complete the isolation of mi RNA in the sample.The results showed that this method has a high recovery efficiency for low-abundance mi RNA,and could be used for mi RNA extraction from serum samples.This provided a feasible sample pre-processing solution for the ultra-rapid detection of mi RNA,and has important research significance for rapid disease diagnosis.