MicroRNAs (miRNAs) are small non-coding RNAs of ∼ 22 nucleotides that are involved in negative regulation of mRNA at the post-transcriptional level. Previously, we developed miRTarBase which provides information about experimentally validated miRNA-target interactions (MTIs). Here, we describe an updated database containing 422 517 curated MTIs from 4076 miRNAs and 23 054 target genes collected from over 8500 articles. The number of MTIs curated by strong evidence has increased ∼1.4-fold since the last update in 2016. In this updated version, target sites validated by reporter assay that are available in the literature can be downloaded. The target site sequence can extract new features for analysis via a machine learning approach which can help to evaluate the performance of miRNA-target prediction tools. Furthermore, different ways of browsing enhance user browsing specific MTIs. With these improvements, miRTarBase serves as more comprehensively annotated, experimentally validated miRNA-target interactions databases in the field of miRNA related research. miRTarBase is available at http://miRTarBase.mbc.nctu.edu.tw/.

MicroRNAs (miRNAs) are small non-coding RNAs of approximately 22 nucleotides, which negatively regulate the gene expression at the post-transcriptional level. This study describes an update of the miRTarBase (http://miRTarBase.mbc.nctu.edu.tw/) that provides information about experimentally validated miRNA-target interactions (MTIs). The latest update of the miRTarBase expanded it to identify systematically Argonaute-miRNA-RNA interactions from 138 crosslinking and immunoprecipitation sequencing (CLIP-seq) data sets that were generated by 21 independent studies. The database contains 4966 articles, 7439 strongly validated MTIs (using reporter assays or western blots) and 348 007 MTIs from CLIP-seq. The number of MTIs in the miRTarBase has increased around 7-fold since the 2014 miRTarBase update. The miRNA and gene expression profiles from The Cancer Genome Atlas (TCGA) are integrated to provide an effective overview of this exponential growth in the miRNA experimental data. These improvements make the miRTarBase one of the more comprehensively annotated, experimentally validated miRNA-target interactions databases and motivate additional miRNA research efforts.

Reverse genetics is a powerful tool to study single-stranded RNA viruses. Despite tremendous efforts having been made to improve the methodology for constructing flavivirus cDNAs, the cause of toxicity of flavivirus cDNAs in bacteria remains unknown. Here we performed mutational analysis studies to identify Escherichia coli promoter (ECP) sequences within nucleotides (nt) 1 to 3000 of the dengue virus type 2 (DENV2) and Japanese encephalitis virus (JEV) genomes. Eight and four active ECPs were demonstrated within nt 1 to 3000 of the DENV2 and JEV genomes, respectively, using fusion constructs containing DENV2 or JEV segments and empty vector reporter gene Renilla luciferase. The Flavivirus genus consists of more than 70 members that are categorized into several antigenic groups (46). Most flaviviruses are transmitted by mosquito or tick vectors and cause serious human and animal diseases (46). They include dengue virus (DENV), Japanese encephalitis virus (JEV), West Nile virus (WNV), yellow fever virus (YFV), and tick-borne encephalitis virus (TBEV). DENV and JEV cause some of the most serious arthropod-borne viral illnesses. There are four different serotypes of dengue virus, DENV1, DENV2, DENV3, and DENV4. Dengue cases have been reported in over 100 countries, and an estimated 2.5 billion people live in areas in which dengue is epidemic (26, 27, 49). DENV infection often leads to dengue fever, dengue hemorrhagic fever, and dengue shock syndrome (24,28,48). JEV transmission has been observed in the Southern Hemisphere and has the potential to become a worldwide public health threat. JEV can cause permanent neuropsychiatric sequelae and is sometimes fatal in children (56,60,61).Flaviviruses are enveloped RNA viruses that consist of single-stranded, positive-sense, 10.5-to 11-kb genomic RNA. The genome is associated with multiple copies of capsid proteins that are translated as a single polyprotein. After entering a host cell, the translated polyprotein is then cleaved into three structural proteins (C, prM, and E) and seven nonstructural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) by host proteases and a single virus-encoded protease to initiate viral replication (12,18). Introduction of flavivirus genomic RNA into susceptible cell lines can result in the production of infectious virus particles (1). This phenomenon has prompted the study of flavivirus virology via introduction of flavivirus genomic RNA that has been transcribed in vitro from fulllength flavivirus infectious cDNA.Reverse genetics is a powerful method for studying the viral replication of positive-strand RNA viruses (8). Unfortunately, the instability of full-length flavivirus cDNA in Escherichia coli has been a major hurdle in a attempt to construct flavivirus cDNAs (reviewed in references 4, 54, and 63). Several strategies have been developed to avoid or overcome the instability of infectious flavivirus cDNA. The instability of plasmids containing full-length YFV was avoided using an in vitro ligation approach involving two plasmids (5...