Comment[ArrayExpressAccession] E-GEOD-48584 MAGE-TAB Version 1.1 Public Release Date 2014-07-02 Investigation Title A comprehensive survey of degradation and 3′ end modification of plant microRNAs by deep sequencing Comment[Submitted Name] A comprehensive survey of degradation and 3′ end modification of plant microRNAs by deep sequencing Experiment Description The degradation and 3′ end modification of plant microRNAs (miRNAs) play crucial roles in regulating miRNA function and stability. However, the process and mechanism of miRNA degradation and 3′ end modification has, to date, been poorly characterized. Here, we report that analysis of the two small RNA libraries constructed from two hickory floral differentiation stages by deep sequencing obtained a large number of truncated miRNAs and miRNAs with 3′ end modifications. The presence of so many truncated miRNAs suggests that plant miRNAs may be degraded through the 5′ to 3′ and 3′ to 5′ ends simultaneously, but the probability of miRNAs being truncated from the 3′ end was higher than from the 5′ end. Single- or double-nucleotide 3′ additions to miRNAs has been observed in many families. In this study, the 3′ addition of adenine to miRNA was the most common, accounting for more than 50% of all miRNA 3′ end modification in both small RNA libraries, followed by uridine addition. This suggests that the 3′ end modification of miRNAs shows a bias towards adenine and uridine in plants. Furthermore, we observed that both truncated miRNA and isomiR expressions associated with mature miRNAs. Our study provides more information regarding the degradation and 3′ end modification of miRNAs in plants. Examination of 2 different female flower buds Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name WANG wang wang Person First Name ZHENGJIA zhengjia zhengjia Person Email wzhj21@163.com Person Affiliation ZHEJIANG A & F UNIVERSITY Person Address ZHEJIANG A & F UNIVERSITY, 88 NORTH CIRCLE ROAD, LIN AN, China Person Roles submitter Protocol Name P-GSE48584-2 P-GSE48584-1 Protocol Description Small clean RNAs were mapped to the genome of hickory sequenced by 454 (unpublished data) and to Populus trichocarpa genome (http://genome.jgi-psf.org/Poptr1_1/) using SOAP software Perfect sRNA matches were retained for further analysis. Sequences matching noncoding RNAs included rRNAs, tRNAs, snRNAs, and snoRNAs in the Rfam (http://www.sanger.ac.uk/Software/Rfam) and NCBI GenBank databases (http://www.ncbi.nlm.nih.gov/blast/Blast.cgi) were discarded. Conserved miRNAs were obtained by BLAST searches, allowing two mismatches, of miRBase 17.0 (http://www.mirbase.org/index.shtml) and were confirmed by predicting the characteristic secondary structures of miRNAs using RNAfold (http://www.tbi.univie.ac.at/~ivo/RNA/ViennaRNA-1.8.1.tar.gz). Mireap software (https://sourceforge.net/projects/mireap/) was used to predict novel miRNAs from the unknown sRNAs in our sequenced genome of hickory. Concentrating on the selected miRNA sequences, we found that some sRNAs perfectly matched the selected miRNA sequences and were trimmed one or more nucleotides at the 3′ or 5′ ends; these were considered to be truncated miRNAs, the intermediates of miRNA degradation. Moreover, we observed that one or more nucleotides at 3′ or 5′ ends, based on the precursor sequences, were added to the sRNAs that perfectly matched the miRNA sequences. The degradation products of these sRNAs were either mature or truncated miRNAs. Therefore, these sRNAs were also used to analyze the degradation of miRNAs. Modifications of miRNA sequences by 3′ nucleotide additions were called isomiRs. We found that to the sRNAs that perfectly matched mature miRNAs were added one or more nucleotides at the 3′ end; these were considered to be isomiRs. There were two kinds of isomiRs, nontemplated and templated, depending on whether or not they were based on the sequences of miRNA precursors. These isomiRs were often appended with A, G, C, or U (e.g., miR156b-A or isomiR-A). small RNA libraries were prepared for sequencing using standard Illumina protocols Protocol Type normalization data transformation protocol nucleic acid library construction protocol Comment[SecondaryAccession] GSE48584 Comment[GEOReleaseDate] 2014-07-02 Comment[ArrayExpressSubmissionDate] 2013-07-08 Comment[GEOLastUpdateDate] 2014-07-02 Comment[AEExperimentType] RNA-seq of non coding RNA Comment[SecondaryAccession] SRP026607 Comment[SequenceDataURI] http://www.ebi.ac.uk/ena/data/view/SRR926847-SRR926848 SDRF File E-GEOD-48584.sdrf.txt