micrornas ( mirnas ) are endogenously encoded small rnas that can regulate gene expressions by base - pairing to protein - coding mrnas for degradation or translation repression . numerous mirnas have been identified from genomes of many animals and plants , such as fruit fly , nematode , zebrafish , chicken , mouse , human , arabidopsis , rice and maize . mirna genes are abundant in humans , estimated to account for 1% of the total predicted genes . in arabidopsis , at least 43 distinct mirna families consisting of 111 members have been reported and archived in although the function of most mirnas remains unknown , a number of mirnas have been shown to play important roles in developmental timing , cell death , cell proliferation , hematopoiesis and patterning of the nervous system in animals , and stress responses , and leaf and flower development in plants ( 26 ) . target duplex free energy may be important for animal mirna target prediction ( 7,8 ) , plant mirna targets can be predicted by sequence similarity since plant mirna seems to bind almost perfectly to its cognate mrna ( 7,9 ) . computational tools have been developed to predict plant mirna targets ( 911 ) , but none is in the web server format . ( 9 ) used patscan ( 12 ) to predict plant mirna targets with 3 mismatches . jones - rhoades and bartel ( 11 ) used their own unpublished programs , together with patscan , and the prediction seems to be more comprehensive . waterman algorithm in mirna target prediction , but failed to detect all previously identified targets ( 10 ) . since most biology laboratories involved in plant mirna research may not have necessary bioinformatic resources for target prediction , a publicly accessible web application for plant mirna target prediction has been developed . the tool allows systematic search for mirna complementary targets in any plant whose genome sequence or a large number of expressed sequence tags ( ests ) are available . backed by an exhaustive search algorithm , the tool is able to find all potential targets with the given mismatches . false positives are reduced by limiting the number of mismatches and by ensuring the target complementarity conservation in another plant species ( 11 ) . the server has a user - friendly and intuitive input interface , as shown in figure 1 . although mirnas are usually 2124 nt ( 4 ) , the input sequence can be in the range of 1928 nt in length to accommodate an sirna input , as the tool can also be used to search for sirna targets and off - targets . to predict target genes , currently , the system includes genome mrnas or ests and other transcripts - assembled gene indices ( 13 ) for 28 plant species downloaded from the institute for genome research ( tigr ) at . with the above input information , a perl script at the backend will then do an exhaustive sequence similarity search , using an algorithm modified from blast ( 14 ) ( see additional file 1 ) . to reduce false positives in predicted targets , the user can limit the number of mismatches , which are classified into three types and are assigned different scores ; the higher scores are for more detrimental mismatches for mirna function : g : u wobble pairings ( each assigned 0.5 scores ) , insertions / deletions ( indels ) ( 2.0 ) and all other non - canonical watson  crick pairings ( 1.0 ) . the total score for an alignment is calculated based on 20 nt . when the query is longer than 20 nt , scores for all possible consecutive 20 nt subsequences are computed and the minimum score is output as the total score for the query - subject alignment . since target complementarity to the mirna 5 end seems to be critical to the target site function ( 1518 ) , any mismatch other than g : u wobble in positions 27 at the 5 end based on the observation that both mirnas and their target sites are evolutionarily conserved across genomes ( 1820 ) , the conservation of target complementarity in another genome can be used to further reduce false positives in plant mirna target prediction ( 11 ) . furthermore , such analysis will also provide useful information about conserved regulatory roles of homologous mirnas in different species . to use this strategy in the server , the information of the homologous mirna and the mrna dataset of the second genome should be provided for the system to do another search . then the system compares potential targets to find whether homologous genes are predicted to be targeted by the homologous mirnas in both genomes . genes are considered to be homologous if they share 1 pfam domains ( 21 ) . all mrna datasets are preprocessed by aligning to pfam - a seed domain sequences ( pfam 16.0 , which contains 7677 families , available at ) . for arabidopsis and rice genome mrna datasets , the corresponding protein datasets are used for functional domain identification using hmmer ( 22 ) with e - value 0.1 as the significance level . since hmmer does not allow dna protein comparisons , all gene index datasets are searched against pfam - a seed domain sequences using blastx program ( 14 ) with e - value cut - off of 10 . tigr 's  eukaryotic gene orthologs dataset ( 23 ) is also used for determining homology relationships in the gene index datasets . the search results are parsed and stored in a mysql database to facilitate the comparisons of target conservation in any two genomes . the first part is a summary of search input parameters , including the query sequence , mismatches allowed and target dataset . the next section is a list of predicted complementary targets displayed in the order of mismatch scores . information shown for each predicted target includes gene identifier , target site position , mismatch score , number of mismatches and target complementary sequence with mismatches highlighted in colors ( green for gu mismatches , purple for indels and red for all other mismatches ) . therefore , the target list includes conserved targets that are highly likely to be true targets . some of these targets may still be true targets since the dataset to be compared for most plants are ests sampled from the genomes and may miss the conserved targets . the last part of the output is the target gene sequences in fasta format , which includes the definition line for the original functional annotation . the target site in the gene can easily be located as it is highlighted in colors ( figure 2 ) . to verify the tool , the prediction of arabidopsis mirna targets by jones - rhoades and bartel ( 11 ) seems to be highly reliable since more than half of the predicted targets were experimentally verified as true targets . in this work , arabidopsis mirnas conserved in rice , as listed in supplementary table s1 in jones - rhoades and bartel ( 11 ) , were used as queries for the tool to predict target genes and the result can be found in additional file 2 . recently , sunkar and zhu ( 6 ) identified stress - regulated mirnas from arabidopsis . they also predicted the potential target genes for these mirnas using the criteria modified from rhoades et al . moreover , the result indicates that sunkar and zhu 's prediction seems to be incomplete . for example , a total of 23 targets were predicted by sunkar and zhu for ten mirnas identified in their experiment , while this server predicts 203 potential targets in total ( see additional file 3 ) . the server aims at predicting plant mirna targets with the highest sensitivity and selectivity by using a search algorithm which guarantees finding all homologous sequences within given mismatches , and by applying current knowledge about mirna targets to minimize false positives . as a practical tool
micrornas ( mirnas ) play important roles in gene expression regulation in animals and plants . since plant mirnas recognize their target mrnas by near - perfect base pairing , computational sequence similarity search can be used to identify potential targets . 
 a web - based integrated computing system , miru , has been developed for plant mirna target gene prediction in any plant , if a large number of sequences are available . 
 given a mature mirna sequence from a plant species , the system thoroughly searches for potential complementary target sites with mismatches tolerable in mirna  target recognition . 
 true or false positives are estimated based on the number and type of mismatches in the target site , and on the evolutionary conservation of target complementarity in another genome which can be selected according to mirna conservation . 
 the output for predicted targets , ordered by mismatch scores , includes complementary sequences with mismatches highlighted in colors , original gene sequences and associated functional annotations . 
 the miru web server is available at .