mature microrna ( mirna ) molecules are approximately 22-nucleotide - long single - stranded rnas that generally repress the expression of protein coding genes .
specifically , they preferentially bind to 3 untranslated regions ( utrs ) of messenger rnas ( mrnas ) and interfere with their stability and translational efficiency ( 1,2 ) .
the first mirnas and their target genes were identified via classical forward genetic techniques in 1993 , but it was not until 2001 that many more mirnas were discovered experimentally and found to be abundant and widespread ( 35 ) . since
then there has been a dramatic growth in the number of annotated mirnas ( figure 1 ) . 

figure 1.the growth of the human mirna genes in mirbase database and the growth of the human experimentally determined mirna target interactions in tarbase .
the growth of the human mirna genes in mirbase database and the growth of the human experimentally determined mirna target interactions in tarbase .
a crucial aspect of the functional analysis of mirnas is the annotation of their protein - coding targets .
a number of computational algorithms have been developed for the prediction of such targets ( 6 ) .
although these programs are very important to guide wet lab experiments , they still lack in sensitivity and specificity ( 7,8 ) . in parallel , and as support for these programs , a number of experimental procedures have been developed to provide indirect or direct support for predicted mirna 
target interactions and results from a growing number of such experiments have been published ( figure 1 ) .
the need for a systematic documentation of such experimentally supported targets was covered by the first version of tarbase ( 9 ) . in 2006 ,
the database recorded over 550 entries with mirna  target interactions in human , mouse , fruit fly , worm and zebrafish . here
we present a substantially updated and extended version of this database , tarbase5.0 , documenting over 1300 entries .
the new database now contains all of the information included in the previous version , plus : 
 specific cells lines ( if any ) used in the experiments , cell - type - specific expression of the gene product and its potential involvement in carcinogenesis , differential expression of mirnas in specific tissues , developmental or pathological events that a specific mirna is involved in and any annotated types of mirna - related mis - regulation in those events , hgnc symbols as defined by hugo ( in order to provide consistent gene naming ) . 

additionally , both the underlying sql database and the user interface have been extensively redesigned with several added external links such as a direct link to the paper 's abstract on the pubmed site .
specific cells lines ( if any ) used in the experiments , cell - type - specific expression of the gene product and its potential involvement in carcinogenesis , differential expression of mirnas in specific tissues , developmental or pathological events that a specific mirna is involved in and any annotated types of mirna - related mis - regulation in those events , hgnc symbols as defined by hugo ( in order to provide consistent gene naming ) . the tarbase5.0 database can be directly accessed from the http://microrna.gr/tarbase web page .
tarbase5.0 contains data extracted from a total of 203 scientific papers resulting in 1333 entries describing a regulatory interaction between a mirna and a target 3 utr ( summarized in table 1 ) . 

table 1.a list of all tarbase5.0 entriesorganismnumber of papersnumber of entriesmicroarray datapsilac datahomo sapiens110285328474mus musculus2810513d .
elegans1814plants2130danio rerio11rat22total203514341474 a list of all tarbase5.0 entries the tarbase5.0 data set contains mirna targets that tested either positive ( induces target gene repression ) or negative ( no influence on target gene expression ) . for each experiment with a positive outcome
the target site is described by the mirna that binds it , the gene in which it occurs , the nature of the experiments that were conducted to test it , the sufficiency of the site to induce translational repression and/or cleavage , and the paper from which all these data is extracted .
additionally , for each mirna and protein - coding gene , the database contains links to several other relevant and useful databases such as ensembl ( 10 ) , hugo ( 11 ) , ucsc genome browser ( 12 ) and swissprot ( 13 ) .
there are a number of direct and indirect experimental procedures that have been developed to test a possible mirna 
the entries in tarbase5.0 are classified into four categories : true or false in the cases where an assay provides direct experimental evidence , or microarray and/or psilac in the cases that present only indirect evidence from high - throughput techniques to measure mirna - mediated global transcriptomic or proteomic changes .
to overexpress a mirna , expression constructs can be engineered using the mature mirna , the precursor ( hairpin ) mirna , or the pri - mirna sequence for transfection into in vitro or in vivo transformed cells .
also , silencing of a specific mirna can be accomplished by introducing chemically modified oligonucleotides that are perfectly complimentary to the mature mirna ( antagomirs ) ( 14 ) .
these methods for modifying mirna expression allow for several types of follow - up techniques to quantify and interpret differences in target gene expression . below we provide a more detailed description of each of the four categories : true or false : the most commonly used method for providing direct experimental evidence is the reporter gene assay . in its simplest form , an expression vector containing a reporter gene [ i.e. luciferase or green fluorescent protein ( gfp ) ] is first modified by cloning the predicted target 3utr downstream of the reporter gene , and then transfected into a cell line of interest in the absence and presence of the cognate mirna . despite the general utility of this approach to assay for 3utr - mediated effects on reporter protein expression , it is not informative for the precise location of the mirna response element ( mre ) or number of mirna target sites in the 3utr .
integration of the reporter gene assay with site directed mutagenesis of the predicted mre ( and , further , restoring the complementarity of the mirna 
mre interaction by mutating the mature mirna sequence ) yields a much more specific and direct result .
to measure effects on reporter mrna levels , the most commonly applied technique is quantitative rt - pcr ( qrt - pcr ) .
measuring effects on both protein and mrna levels can help provide information about the mode of mirna - mediated silencing : mrna translational repression or immediate risc - mediated mrna cleavage and degradation . a mirna 
mre interaction is reported as true or false based on the results of the reporter gene assay .
microarray and/or psilac : these high - throughput approaches measure global changes in the transcriptome ( 15 ) or proteome ( 8,16 ) given the presence or absence of a mirna . despite their power for large - scale analysis , these techniques only provide indirect evidence about a mirna 's targets since it is not possible to distinguish between primary direct targets and secondary indirect targets . other high - throughput methods like degradome sequencing ( 17,18 ) are also immensely useful but only in the scenarios where a mirna induces risc - mediated mrna cleavage . in order to facilitate user interaction , the query function is divided into several functionally related subgroups .
the initial screen of the tarbase5.0 user interface allows users to query based on mirna , gene and organism . for more advanced queries ,
the user can utilize the extended query options . in this case , the search menus are arranged into four functionally related groups .
target interaction : the validity of the interaction ( field  support type , either true or false ) , the function of the interaction which can be either translational repression or mrna cleavage ( field  datatype ) , the sufficiency of a single target site to exert the specific function ( field  s_s_s ) and the number of mirna response elements present in the specific utr ( field  mre ) .
the second group contains the fields that refer to the experimental methods that led to the reported result .
target interaction ( i.e. reporter gene assays ) while  indirect support refers to experimental procedures that provide more global , system - wide mirna - mediated effects ( i.e. microarrays ) .
the third group corresponds to biological properties of the mirna or target gene : biological functions ( field  protein type ) , specific expression profiles ( field  mirna expression ) or the physiological processes in which this interaction is involved ( field  event or pathology ) .
the fourth and final group contains some general query features such as the scientific paper ( searchable by author or pmid ) .
, the results screen ( figure 2 ) shows only the repression type , the mirna identifier , the target gene identified by the hgnc symbol ( if it is a human gene ) , the common gene name , the refseq isoform i d ( particularly relevant in cases of gene variants or snp haplotypes ) , the affected biological processes and the paper containing the information presented . users can opt to view more detailed information by clicking on the 
+  box so that the expanded results view is opened ( figure 2 ) .
the additional information is divided into three categories : mirna information , gene information and experimental conditions . 

the  mirna information category contains the properties of the specific mirna such as the mirna 's sequence [ extracted from mirbase ( 19 ) ] , the number and sequences of the mres , their locations within the gene 's 3utr , and the affected tissues ( extracted from the paper ) .
the  gene information category gathers mostly biological properties of the target gene like the protein type , ensembl and swissprot ids and chromosome location , providing direct links to ensembl , swissprot and the ucsc browser respectively .
moreover , expression profiles and tumor involvement are also provided for human genes ( information extracted from the ensembl egenetics database ) .
finally , the  experimental conditions category provides the nature of the direct or indirect evidence for the mirna  target gene interaction .
the cell lines used to carry out the specific experiment are also presented in order to render the experimental conditions more complete and reproducible .
even though several computational programs exist to predict mirna targets , the necessity for a systematic collection and description of mirna targets with experimental support led to the development of tarbase .
the continuously expanding number of known and newly identified mirnas and their targets , combined with their central role in biological systems , renders this field particularly dependent on centralized information that is accurate , up - to - date , comprehensive and easy to browse or download . in order to satisfy these requirements
, we have made extensive updates and modifications and present the new version of the database , tarbase5.0 .
the tarbase data files can be freely downloaded and used according to the gnu public license .
funding for open access charge : aristeia award from general secretary research and technology , greece .
 tarbase5.0 is a database which houses a manually curated collection of experimentally supported microrna ( mirna ) targets in several animal species of central scientific interest , plants and viruses . 
 mirnas are small non - coding rna molecules that exhibit an inhibitory effect on gene expression , interfering with the stability and translational efficiency of the targeted mature messenger rnas . even though several computational programs exist to predict mirna targets , 
 there is a need for a comprehensive collection and description of mirna targets with experimental support . 
 here we introduce a substantially extended version of this resource . 
 the current version includes more than 1300 experimentally supported targets . 
 each target site is described by the mirna that binds it , the gene in which it occurs , the nature of the experiments that were conducted to test it , the sufficiency of the site to induce translational repression and/or cleavage , and the paper from which all these data were extracted . 
 additionally , the database is functionally linked to several other relevant and useful databases such as ensembl , hugo , ucsc and swissprot . 
 the tarbase5.0 database can be queried or downloaded from http://microrna.gr/tarbase . 
INTRODUCTION
METHOD AND RESULTS
CONCLUSION
AVAILABILITY
FUNDING