siRNA is a double-stranded RNA containing 21 nucleotides (nt), with high sequence specificity. After cell transfection, siRNA can trigger rapid degradation of the corresponding mRNA in a short duration. shRNA (short hairpin RNA) is an RNA sequence of tight hairpin loop, including two short reverse repeats and a loop region connecting the two repeats. shRNA can be cloned into a plasmid vector as a "short hairpin" structure. After plasmid insertion, hairpin sequence is expressed to form double-stranded RNA (dsRNA) and enters the RNAi pathway to achieve gene silencing. The final goal of gene silencing is achieved through the RNA interference (RNAi) pathway. The expression of shRNA in cells can be achieved in vitro by transferring plasmids, viral or bacterial vectors. Among these vectors, virus-infected cells contain high efficiency in vivo, which solves the problem of low plasmid transfection efficiency for certain cell types (including primary cells and less dividing cells).
Fig.1 Differences in siRNA(left) and shRNA(right) mediated RNA interference pathways. ( Rao D, 2009)
Design and prepare 4 shRNA vectors for the target genes, ensuring that at least one shRNAs has a suppression efficiency of 70% or more at the mRNA level.
|shRNA for target genes||4|
|positive control shRNA vector||1|
|negative control shRNA vector||1|
We can insert the DNA encoding your target shRNA into the plasmid and perform sequence correctness testing. Simply inform us the target gene sequence or Gene ID number, and name of the vector you want to insert (such as pRS, pGFP-V-RS or pRFP-C-RS) for modeling. Nevertheless, we provide enough usable, purified expression plasmids to encode your target shRNA.
After cloning siRNA fragments into target expression vectors, customers need to prepare a large number of high-purity or endotoxin-free plasmids for the next experiment. BOC Sciences offers a service for the preparation of high-purity plasmids.
1) Provide plasmids to be prepared (>20 ng).
2) Plasmid resistance and macrorestriction map.