Gene knockdown by RNAi in the pea aphid <Emphasis Type="Italic">Acyrthosiphon pisum</Emphasis>

Background  

RNA interference (RNAi) is a powerful method to inhibit gene expression in a sequence specific manner.     

Functional complementation of RNA interference mutants in trypanosomes

Background  

In many eukaryotic cells, double-stranded RNA (dsRNA) triggers RNA interference (RNAi), the specific degradation of RNA of homologous sequence. RNAi is now a major tool for reverse-genetics projects, including large-scale high-throughput screens. Recent reports have questioned the specificity of RNAi, raising problems in interpretation of RNAi-based experiments.     

Comparison of chicken 7SK and U6 RNA polymerase III promoters for short hairpin RNA expression

Background  

RNA polymerase III (pol III) type 3 promoters such as U6 or 7SK are commonly used to express short-hairpin RNA (shRNA) effectors for RNA interference (RNAi). To extend the use of RNAi for studies of development using the chicken as a model system, we have developed a system for expressing shRNAs using the chicken 7SK (ch7SK) promoter.     

Gene silencing by RNAi in mouse Sertoli cells

Background  

RNA interference (RNAi) is a valuable tool in the investigation of gene function. The purpose of this study was to examine the availability, target cell types and efficiency of RNAi in the mouse seminiferous epithelium.     

Criteria for effective design, construction, and gene knockdown by shRNA vectors

Background  

RNA interference (RNAi) technology is a powerful methodology recently developed for the specific knockdown of targeted genes. RNAi is most commonly achieved either transiently by transfection of small interfering (si) RNA oligonucleotides, or stably using short hairpin (sh) RNA expressed from a DNA vector or virus. Much controversy has surrounded the development of rules for the design of effective siRNA oligonucleotides; and whether these rules apply to shRNA is not well characterized.     

Acetylation of PAMAM dendrimers for cellular delivery of siRNA

Background  

The advancement of gene silencing via RNA interference is limited by the lack of effective short interfering RNA (siRNA) delivery vectors. Rational design of polymeric carriers has been complicated by the fact that most chemical modifications affect multiple aspects of the delivery process. In this work, the extent of primary amine acetylation of generation 5 poly(amidoamine) (PAMAM) dendrimers was studied as a modification for the delivery of siRNA to U87 malignant glioma cells.     

Influence of RT-qPCR primer position on EGFR interference efficacy in lung cancer cells

Background  

Real-time quantitative RT-PCR (RT-qPCR) is a "gold" standard for measuring steady state mRNA levels in RNA interference assays. The knockdown of the epidermal growth factor receptor (EGFR) gene with eight individual EGFR small interfering RNAs (siRNAs) was estimated by RT-qPCR using three different RT-qPCR primer sets.     

HIV-1 TAR element is processed by Dicer to yield a viral micro-RNA involved in chromatin remodeling of the viral LTR

Background  

RNA interference (RNAi) is a regulatory mechanism conserved in higher eukaryotes. The RNAi pathway generates small interfering RNA (siRNA) or micro RNA (miRNA) from either long double stranded stretches of RNA or RNA hairpins, respectively. The siRNA or miRNA then guides an effector complex to a homologous sequence of mRNA and regulates suppression of gene expression through one of several mechanisms. The suppression of gene expression through these mechanisms serves to regulate endogenous gene expression and protect the cell from foreign nucleic acids. There is growing evidence that many viruses have developed in the context of RNAi and express either a suppressor of RNAi or their own viral miRNA.     

High-throughput isolation of ultra-pure plasmid DNA by a robotic system

Background  

Short hairpin RNA (shRNA) encoded within an expression vector has proven an effective means of harnessing the RNA interference (RNAi) pathway in mammalian cells. A survey of the literature revealed that shRNA vector construction can be hindered by high mutation rates and the ensuing sequencing is often problematic. Current options for constructing shRNA vectors include the use of annealed complementary oligonucleotides (74 % of surveyed studies), a PCR approach using hairpin containing primers (22 %) and primer extension of hairpin templates (4 %).     

Characterisation and application of a bovine U6 promoter for expression of short hairpin RNAs

Background  

The use of small interfering RNA (siRNA) molecules in animals to achieve double-stranded RNA-mediated interference (RNAi) has recently emerged as a powerful method of sequence-specific gene knockdown. As DNA-based expression of short hairpin RNA (shRNA) for RNAi may offer some advantages over chemical and in vitro synthesised siRNA, a number of vectors for expression of shRNA have been developed. These often feature polymerase III (pol. III) promoters of either mouse or human origin.     

Identification of clustered microRNAs using an <Emphasis Type="Italic">ab initio</Emphasis> prediction method

Background  

MicroRNAs (miRNAs) are endogenous 21 to 23-nucleotide RNA molecules that regulate protein-coding gene expression in plants and animals via the RNA interference pathway. Hundreds of them have been identified in the last five years and very recent works indicate that their total number is still larger. Therefore miRNAs gene discovery remains an important aspect of understanding this new and still widely unknown regulation mechanism. Bioinformatics approaches have proved to be very useful toward this goal by guiding the experimental investigations.     

In-depth sequencing of the siRNAs associated with peach latent mosaic viroid infection

Background  

It has been observed that following viroid infection, there is an accumulation of viroid-derived siRNAs in infected plants. Some experimental results suggest that these small RNAs may be produced by the plant defense system to protect it from infection, indicating that viroids can elicit the RNA-silencing pathways. The objective of this study is to identify in the peach latent mosaic viroid (PLMVd), a model RNA genome, the regions that are most susceptible to RNA interference machinery.     

sIR: siRNA Information Resource,a web-based tool for siRNA sequence design and analysis and an open access siRNA database

Background  

RNA interference has revolutionized our ability to study the effects of altering the expression of single genes in mammalian (and other) cells through targeted knockdown of gene expression. In this report we describe a web-based computational tool, siRNA Information Resource (sIR), which consists of a new open source database that contains validation information about published siRNA sequences and also provides a user-friendly interface to design and analyze siRNA sequences against a chosen target sequence.     

Suppression of RNA interference increases alphavirus replication and virus-associated mortality in Aedes aegypti mosquitoes

Background  

Arthropod-borne viruses (arboviruses) can persistently infect and cause limited damage to mosquito vectors. RNA interference (RNAi) is a mosquito antiviral response important in restricting RNA virus replication and has been shown to be active against some arboviruses. The goal of this study was to use a recombinant Sindbis virus (SINV; family Togaviridae; genus Alphavirus) that expresses B2 protein of Flock House virus (FHV; family Nodaviridae; genus Alphanodavirus), a protein that inhibits RNAi, to determine the effects of linking arbovirus infection with RNAi inhibition.     

A versatile approach to multiple gene RNA interference using microRNA-based short hairpin RNAs

Background  

Effective and stable knockdown of multiple gene targets by RNA interference is often necessary to overcome isoform redundancy, but it remains a technical challenge when working with intractable cell systems.