Team:AMU-Poznan/Project
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sh-miR Designer v2.0Sh-miR designer is a project we started during iGEM 2013. This year we would like to continue and expand functionality of the software. sh-miR designer v1.0 (link) is aimed to create sh-miR molecules based on siRNAs provided by the user. In sh-miR designer v2.0 only the mRNA number (from NCBI database), which expression should be decreased can be provided. Moreover, we expanded functionality of the software with off-target validation and check of immune motifs and also extended miRNA-shuttles database. sh-miR Designer is a software aimed for fast and efficient design of effective RNA interference (RNAi) reagents - sh-miRs, also known as artificial miRNAs. sh-miRs are RNA particles whose structure is based on miRNA precursor pri-miRNA, but sequence interacting with transcript is changed depending on research purpose. Maintenance of structure of pri-miRNA is very important to enable cellular processing and therefore ensure functionality of artificial particles. sh-miRs delivered to cells on genetic vectors - plasmids or viral vectors - enter natural RNAi pathway and silence target mRNA. They can be used in genetic therapies and basic biomedical research. We will provide two applications to access the software, one which require siRNA sequences and the second which require transcript accession number from NCBI database. Each user will receive an account with login (e-mail) and password where he/she would be able to check history of software usage.Standard PartsWe are creating a particle which can be inserted into plasmid built by Standard parts from Registry of Standard Biological Parts.User GuideTechnologyAPI documentationAPI for developersReleases (re-use of 2013iGEM software)sh-miR designer v1.0 First version of the program presented on iGEM 2013 by our team. The user have to provide siRNA sequence (one or both strands). The miRNA database includes 5 miRNAs. sh-miR designer v1.1 Strand discrimination functionality was added based on miRbase. extended miRNA database sh-miR designer v2.0 siRNA prediction algorithm was added to functionality (the input is NCBI transcript number) user can choose GC content of siRNA molecule, maximal off-target, if he/she wants immunostimulatory sequences inside the molecule and to choose if he/she wants to include all or chosen scaffold extended miRNA database What we are working on: strand discrimination based on therodynamics of siRNA ends; preparing construct with flanking restriction enzymes providing plasmids with parts from standard partsUnit TestsValidationTest cases1) ATXN3 - therapeutic sh-miR against SCA3 disease transcript number: NM_001164782.1 GC content: default Off-target: 10 scaffold: ALL immunostimulatory: NO 2) P54 - therapeutic sh-miR against cancer transcript number: NM_000546.5 GC content: default Off-target: 10 scaffold: ALL immunostimulatory: YES 3) biological function analysis, PARP1, we have miR-30a in our lab transcript number: NM_001618.3 GC content: default Off-target: 0 scaffold: miR-30a immunostimulatory: no_difference (default) 4) biological function, PARP1, we have miR-30a in our laboratory and have immune cells transcript number: NM_001618.3 GC content: default Off-target: 0 scaffold: miR-30a immunostimulatory: NO 5) biological function, PARP1 and have immune cells transcript number: NM_001618.3 GC content: default Off-target: 0 scaffold: ALL immunostimulatory: NOReferences1. 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