SSD - a free software for designing multimeric mono-, bi- and trivalent shRNAs

Abstract

RNA interference (RNAi) is a powerful gene silencing technology, widely used in analyses of reverse genetics, development of therapeutic strategies and generation of biotechnological products. Here we present a free software tool for the rational design of RNAi effectors, named siRNA and shRNA designer (SSD). SSD incorporates our previously developed software Strand Analysis to construct template DNAs amenable for the large scale production of mono-, bi-and trivalent multimeric shRNAs, via in vitro rolling circle transcription. We tested SSD by creating a trivalent multimeric shRNA against the vitellogenin gene of Apis mellifera. RT-qPCR analysis revealed that our molecule promoted a decrease in more than 50% of the target mRNA, in a dose-dependent manner, when compared to the control group. Thus, SSD software allows the easy design of multimeric shRNAs, for single or multiple simultaneous knockdowns, which is especially interesting for studies involving large amounts of double-stranded molecules.

A small interfering RNA (siRNA) database for SARS-CoV-2

“…Thus, the proposed database provides information about each possible 18 to 21 nucleotides siRNA target region from SARS-CoV-2, one table for each length. Moreover, tools employed for assessing siRNAs efficiency (20)(21)(22) operate over sequences lying in that range, which reinforces our choice. Since they present the same columns, we explain here the development process only for the 21-length table.…”

“…Sequence properties regarding base context and alignment information were calculated from the above sequences sets and performed alignments (see Supplementary Text 1). Thermodynamic information and expected efficiency of candidates siRNA designed for targeting those regions was calculated with OligoCalc 7 (24) and three predictors, namely ThermoComposition21 (20), SSD (20,22), and si-shRNA Selector (21),…”

A small interfering RNA (siRNA) database for SARS-CoV-2

“…Thus, the proposed database provides information about each possible 18–21 nucleotides siRNA target region from SARS-CoV-2, one table for each length. Moreover, tools employed for assessing siRNAs efficiency 45 , 61 , 62 operate over sequences lying in that range, which reinforces our choice. Since they present the same columns, we explain here the development process only for the 21-length table.…”

“…For example, our proposed database provides information regarding the bases, GC, and AU context, so as the quantities of each RNA nitrogenated base in sequences, besides information about the presence of UUUU and GCCA, considered toxic motifs 41 , so any user with a proper efficacy evaluation method (or anyone provided by literature) can easily evaluate siRNAs with this database at disposal. It also provides thermodynamic information collected from the application of three predictors 45 , 61 , 62 , thus enabling users to have a deeper look at siRNAs’ properties, and choose the best ones according to their specificities. As it can be seen in Fig.…”

“…Sequence properties regarding base context and alignment information were calculated from the above sequences sets and performed alignments (see Supplementary Text S1 ). Thermodynamic information and expected efficiency of candidates siRNA designed for targeting those regions was calculated with OligoCalc 68 and three predictors, namely ThermoComposition21 61 , SSD 62 , and si-shRNA Selector 45 . Finally, we elicited criteria filtering sets from literature regarding (a) Toxicity, (b) Efficiency, (c) Populations and genomes coverages, (d) Stability, and (e) Effectiveness prediction, which are summarized at Table 2 , and then analyzed whether they could be assessed with the features of proposed database.…”