Differential Expression Analysis And Function Analysis Of Tomato MiRNAs Under Botrytis Cinerea Treatment

MicroRNAs (miRNAs) are single-stranded RNA molecules of about22nucleotides inlength that widely exist in plant and animal cells. MiRNAs function as transcriptional orpost-transcriptional regulators through base pairing with target mRNAs.In recent years, a plenty of miRNAs have been identified in many plant species, andmainly in the whole genome sequencing model plant such as Arabidopsis and rice. However,few have been discovered in other species.As an important economic crop, tomato is widely cultivated all over the world. There arecurrently377Arabidopsis miRNAs and713rice miRNAs in miRBase, but only42miRNAsare found in tomato. Visibly, there are still a lot of tomato miRNAs need to be discovered.Botrytis cinerea is a worldwide severe crop disease, and it mainly causes the rotten fruit, fruityield and quality of tomato. Some studies of plant miRNAs resistant adversity stress havebeen reported, but relatively few have been of tomato miRNAs resistant botrytis cinerea.This study uses the next generation of high-throughput sequencing and bioinformaticsanalysis method, combined with the current popular miRNAs prediction algorithm, aiming atovercoming the current miRNAs analysis programs’ drawback such as long operation timeand high requirements for computer hardware, and developing a plant miRNAs annotationand qualification tool that perform plant miRNAs identification and qualification in arelatively short time by a computer with general hardware.Through analysis of the mainstream of miRNAs tools compared with our developmenttools, the tool with high efficiency was selected to perform the identification and qualificationof tomato small RNA libraries. The targets of the differentially expressed miRNAs werepredicted with the help of degradome sequencing library. Finally, the target mRNAs weresent to functional annotation using blast2go, and botrytis cinerea responsive miRNAs wereanalyzed.The results are as follow: 1. We developed a plant miRNAs annotation and qualification tool, miR-island, and itovercome current miRNAs analysis programs’ drawback such as long operation time and highrequirements for computer hardware, and perform the identification and qualification of smallRNA libraries with a general office computer in a relatively short time. MiR-island shows asignificant advantage when comparing with miRDeep-P or ShortStack.2. By analyzing tomato small RNA libraries in two conditions using miR-island package,a total of145miRNAs were expressed. Among that,41are tomato known miRNAs inmiRBase, and104are tomato newly discovered miRNAs.3. By annotation and qualification of two small RNA sequencing libraries, the resultshowed that under botrytis cinerea stress treatment, a total of32non-redundant sequencesrepresented for47miRNAs are differentially expressed. Among them,9non-redundantsequences represented for13tomato known miRNAs and23non-redundant sequencesrepresented for34tomato newly discovered miRNAs. The9non-redundant sequences(represented for13tomato known miRNAs) are up-regulated, while among the left23non-redundant sequences (represented for34newly discovered miRNAs),10non-redundantsequences (represented for12miRNAs) are down-regulated, and the left13non-redundantsequences (represented for22miRNAs) are up-regulated.4. The differentially expressed miRNAs were performed for targets annotation with thehelp of degradome sequencing data using CleaveLand4. There were9differentially expressednon-redundant sequences that have a total of24potential target mRNAs. The function of the4target mRNA of sly-miR482b are all disease resistant protein and it implied thatsly-miR482b plays an important role in responding botrytis cinerea stress treatment.