RNAi Technology Protocols

Protocol describes a procedure to anneal sense and antisense siRNA strands to form a duplex.  The duplexes can then be transfected into cultured cells. - [Read Annealing siRNAs to Produce siRNA Duplexes Protocol]

Protocol describes the preparation of an shRNA-expressing Gateway entry vector.  This process is preceded by the design and synthesis of target gene-specific sense and antisense oligonucleotides which, when annealed, will constitute an shRNA cassette. - [Read Annealing, Cloning, and Sequencing of shRNA Linkers into pEN_H1 Plasmids Protocol]

Protocol for cloning siRNA duplexes into pSUPER or pSUPERretro vectors. - [Read Cloning siRNA Duplexes into pSUPER or pSUPERretro Vectors Protocol]

Information on how detect and quantitate MicroRNA in laser capture microdissection samples. Includes: Obtain Laser Capture Microdissected Samples; Isolate miRNA from LCM Samples; Quantitate miRNA by qRT-PCR; Detection of miRNA in Microdissected Tissue from Mouse Brain by qRT-PCR; Differential Expression of MicroRNA in Whole Brain Tissue Compared to a More Homogeneous Population of Cells. - [Read Detect and Quantitate MicroRNA in Laser Capture Microdissection Samples]

Protocol for electroporation and RNAi in the rodent retina in vivo and in vitro. - [Read Electroporation and RNAi in the Rodent Retina In Vivo and In Vitro Protocol]

Short interfering RNAs (siRNAs) can be used to prime RNA synthesis by the RNA-dependent RNA polymerase (RdRP).  SiRNAs can be used by RdRP as primers for specific cellular mRNAs, forming dsRNA products capable of inducing transitive RNAi. - [Read Protocol for siRNA-Primed RNA Synthesis Protocol]

Protocol for si-RNA transfection of mouse bone marrow progenitors by electroporation. - [Read si-RNA Transfection of Mouse Bone Marrow Progenitors by Electroporation Protocol]

Protocol Involves the transfection of siRNA into RAW 264. 7 cells using Lipofectamine 2000.  Cells are transfected with siRNA twice (on subsequent days).  Target gene knockdown is assessed from total RNA isolated 48 hr post-transfection or from protein isolated 72 hr post-transfection. - [Read siRNA Double Transfection of RAW 264.7 Cells with Lipofectamine Protocol]

Protocol Involves the transfection of siRNA into RAW 264. 7 cells using Lipofectamine 2000.  Cells are transfected with siRNA twice (on subsequent days).  Target gene knockdown is assessed from total RNA isolated 48 hr post-transfection or from protein isolated 72 hr post-transfection. - [Read siRNA Double Transfection of RAW 264.7 Cells with Lipofectamine Protocol II]

The siRNA user guide. Includes: Selection of siRNA duplexes from the target mRNA sequence; Preparation of siRNA duplexes; Useful siRNA reagent combinations; Transfection of siRNA duplexes; Sequences of siRNA duplexes used in our studies. - [Read The siRNA User Guide]