Discover Structural Noncoding RNA Using Comparative Genomics And Biochemical Methods

Noncoding RNA refers to RNA that does not encode protein,including miRNA,lncRNA,ribozyme,riboswitch,etc.they usually undertake the important gene regulation function in organisms,and link with diseases.Therefore,noncoding RNA has gradually become a research hotspot in the biological community.This project will work in three directions: 1.Discovery of algae structural ncRNA by improved bioinformatics pipeline(CM line);2.Discovery of variants of algae ribozymes and riboswitches by Infernal;3.Establishment of a new ribozyme discovery method based on the specificity of ribozyme substrate by high-throughput sequencing.There are two main methods to find more structural noncoding RNAs: Experimental and bioinformatics methods.Using experimental method to find more ncRNAs was usually focused on the ncRNAs which are well characterized.With the completion of genome sequencing and the establishment of various databases,it is possible and necessary to discover and predict ncRNA on a large scale.With the rapid development of the bioinformatics,a large number of bioinformatics tools have been developed,which can be applied to the search of noncoding RNAs effectively.In this study,I used the improved bioinformatics pipeline(CM line)to discover the structural ncRNA of algae by using the existing bioinformatics tools,clustering the noncoding regions of algae genome,predicting the secondary structure.I found a large number of new structured noncoding RNA.According to the secondary structure characteristics,there are 47589 motifs with more than 80 scores,including those that may control the variable splicing process by binding with upstream and downstream polypeptide molecules.The variants of algal ribozymes and riboswitches were found by Infranal,and the secondary structural descriptions of ribozymes and riboswitches were from Rfam,which is a non coding RNA database.The self cleaving activity of HDV and hammerhead ribozymes in algal genomes was verified by designed experiments..Based on the specificity of ribozyme substrate,a new ribozyme method was established by using chemical modification and high-throughput sequencing.Using the characteristics of natural ribozyme self cutting in vivo,the 5 '-OH terminal was modified.Through the innovative design of experimental scheme,ribozyme cutting products was enriched.The candidate sequences of 8 natural ribozyme cutting products in Bacillus subtilis genome were obtained by high-throughput sequencing technology.And I am designing experiments to verify the self cleaving activity of the candidate sequences.Through the above research,we have established different methods to discover structured noncoding RNA,have found some new structured noncoding RNA,variants and have also found known ncRNAs in new species,also explored the biological functions of some ncRNAs,and provided new methods and experience for the research of noncoding RNAs.