knockdown Protocols

Category: Transfection Protocols: Stable Transfection Protocols

The AfCS is utilizing antisense technology to manipulate signaling protein expression in the RAW 264.7 macrophage-like cell line. This can be achieved by the transfection of gene-specific antisense oligonucleotides (ASOs). The following procedure involves the transfection of ASOs into RAW 264.7 cells using FuGENE 6 transfection reagent. Subsequently, the isolated total RNA or protein from these transfected cells can be used to assess the level of mRNA or protein knockdown, respectively. - [Read Antisense Oligonucleotide Transfection of RAW 264.7 Cells with FuGENE 6 in a 24-Well Dish]

Category: RNAi Technology Protocols: RNAi Mouse Embryos and Oocytes Protocols

Protocol describes a method to collect early embryos from 6-week-old mice.  Subsequently, the isolated embryos can be injected with double-stranded RNA to induce knockdown of a gene of interest. - [Read Collection of Early Mouse Embryos for RNAi Protocol]

Category: RNAi Technology Protocols: RNAi Mouse Embryos and Oocytes Protocols

Protocol describes a method to collect oocytes from 6-week-old mice.  Subsequently, the isolated oocytes can be injected with double-stranded RNA to induce knockdown of a gene of interest. - [Read Collection of Mouse Oocytes for RNAi Protocol]

Category: RNAi Technology Protocols: RNAi Mammalian Cells Protocols

Protocol describes the use of western blotting to analyze the RNAi-induced depletion of target protein from mammalian cells.  Immunofluorescence can be used as an alternative. - [Read Detection of RNAi-Induced Protein Knockdown in Mammalian Cells by Western Blotting Protocol]

Category: Western Blot Protocols

Protocol describes the use of western blotting to analyze the RNAi-induced depletion of target protein from mammalian cells.  Immunofluorescence can be used as an alternative. - [Read Detection of RNAi-Induced Protein Knockdown in Mammalian Cells by Western Blotting Protocol]

Category: Immunology: T Cell Protocols

Protocol for Dlgh1 knockdown in primary OT-1 T lymphocytes. - [Read Dlgh1 Knockdown in Primary OT-1 T Lymphocytes Protocol]

Category: Transfection Protocols

Transfection protocol used in the Dlgh1 knockdown in primary OT-1 T lymphocytes. - [Read Dlgh1 Knockdown in Primary OT-1 T Lymphocytes: Transfection Protocol]

Category: Immunology: Immunofluorescence Protocols

This protocol describes the use of a specific antibody that recognizes the targeted gene product to detect RNAi-induced gene knockdown in mammalian cells. Western blot technology can be used as an alternative (see Detection of RNAi-Induced Protein Knockdown in Mammalian Cells by Western Blotting). - [Read Immunofluorescence Detection of RNAi-Induced Protein Knockdown in Mammalian Cells Protocol]

Category: RNAi Technology Protocols: RNAi Mammalian Cells Protocols

Protocol describes the use of a specific antibody that recognizes the targeted gene product to detect RNAi-induced gene knockdown in mammalian cells.  Western blot technology can be used as an alternative. - [Read Immunofluorescence Detection of RNAi-Induced Protein Knockdown in Mammalian Cells Protocol]

Category: Primary Cell Isolation and Culture Protocols: Mammalian Cell Culture Protocols

This protocol introduces a method for maintaining mammalian cell culture suitable for treatment with siRNA. Transfection of cultured cells with siRNAs (see Transfection of siRNA Duplexes into Mammalian Cells) and downstream analysis of the knockdown cells. - [Read Mammalian Cell Culture and Preparation of Cells for RNAi in 24-Well Plates Protocol]

Category: RNAi Technology Protocols: RNAi Mammalian Cells Protocols

Protocol introduces a method for maintaining mammalian cell culture suitable for treatment with siRNA.  Transfection of cultured cells with siRNAs and downstream analysis of the knockdown cells. - [Read Mammalian Cell Culture and Preparation of Cells for RNAi in 24-Well Plates Protocol]

Category: RNAi Technology Protocols: RNAi Mouse Embryos and Oocytes Protocols

Protocol describes the use of RT-PCR to confirm knockdown of a gene of interest in RNAi-treated mouse oocytes and early embryos.  All of the solutions used during RNA isolation and reverse transcription must be Rnase-free - [Read RNA Isolation and RT-PCR from dsRNA-Treated Mouse Oocytes and Early Embryos Protocol]

Category: RNAi Technology Protocols

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]

Category: RNAi Technology Protocols

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]