X-tremeGENE™ siRNA Transfection Reagent
ROCHE/SITRAN-RO - Polymer reagent for delivering siRNA to common cell lines
Product Type: Chemical
Effective and specific gene knockdown of EGFP expression. HEK-293 cells were cotransfected with either an EGFP plasmid (1 1/4g) and a GFP-specific siRNA, or the same amount of EGFP plasmid and a luciferase (Luc)-specific siRNA (non-silencing siRNA control). Cotransfection-based gene-knockdown experiments were analyzed by western blotting. EGFP was detected on the blot with a specific antibody. Results showed specific expression of EGFP. Knockdown was observed only with the GFP-specific siRNA.
Silencing of an expression plasmid by siRNA cotransfection. Human 293T embryonic kidney cells were cotransfected with 0.6 μg expression vector for FLAG-tagged protein and 6.25 μl of siRNA solution (either a 20 μM solution of a specific siRNA or the same concentration of a control siRNA directed against lamC). The transfection procedure was performed using either X-tremeGENE siRNA Transfection Reagent or reagents from three other suppliers. Without any further medium change, the cells were incubated for 24 hours after lipofection. The cells were then viewed under a microscope and their vitality was assessed. Cell proteins were analyzed by immunoblot according to standard methods. The arrow marks the expected position of the FLAG-tagged protein. Equal amounts of cellular proteins (20 μg/lane) were applied to the original gel. As judged by specific repression of protein expression, only two of the reagents, the one from supplier A and X-tremeGENE siRNA Transfection Reagent, were capable of delivering DNA and siRNA simultaneously into the cells. A clear inhibition of target gene expression by specific siRNA was seen after lipofection with either the reagent from supplier A or X-tremeGENE siRNA Transfection Reagent. When the performance of these two reagents was compared, however, X-tremeGENE siRNA Transfection Reagent gives far better results; more protein was expressed after transfection with X-tremeGENE siRNA Transfection Reagent, demonstrating the high transfection efficiency obtained with this compound. By comparison, the other two reagents (B and C) did not perform well. When the reagent from supplier B was used, protein expression was detected, but the RNAi effect was completely absent, indicating that RNA was not efficiently transfected with this compound. Lipofection component from supplier C was completely ineffective in delivering the DNA/RNA mixture.
Comparison of transfection reagent effectiveness in four different cell types. Cells were transfected with siRNA specific for HPRT (hypoxanthine-phosphoribosyl-transferase) using different siRNA transfection reagents (including X-tremeGENE siRNA Transfection Reagent). Transfection was performed according to the suppliers recommended protocol or a standard protocol using 100 nM siRNA. The LightCycler® h-HPRT Housekeeping Gene Set and the LightCycler® System were used to measure knockdown efficiency.
| concentration | 1 mg/mL |
| grade | for molecular biology |
| manufacturer/tradename | Roche |
| packaging | pkg of 1 mL (04476093001 [1 mg/ml]) |
| pkg of 5 × 1 mL (04476115001 [5 x 1 mg/ml]) | |
| Quality Level | 100  |
| shipped in | wet ice |
| storage temp. | 2-8°C |
| technique(s) | transfection: suitable |
| usage | sufficient for 2,000 transfections (04476115001) |
| sufficient for 400 transfections (04476093001) |
| Application: | X-tremeGENE siRNA Transfection Reagent efficiently delivers short interfering RNA (siRNA) into many commonly used cell types including HeLa, NIH 3T3, HEK-293, CHO-K1, and COS-7, and several hard-to-transfect cell lines, such as HT29, a human adenocarcinoma cell line. |
| Features and Benefits: | • Knock down gene expression over 90% in many different cell types. • Maximize experimental flexibility with a single reagent for siRNA- and cotransfection-based gene-knockdown experiments. • Produce meaningful results using a reagent that exhibits low cytotoxicity, ensuring that the cellular effects you observe are due to the transfected siRNA rather than the transfection procedure. • Work with or without serum, avoiding medium changes (e.g., to serum-free medium) before or after transfection.; X-tremeGENE siRNA Transfection Reagent is a proprietary blend of lipids and other components, free of animal products. |
| General description: | Short interfering RNA (siRNA) can be directly introduced into cells by transfection. A lipid-based transfection reagent, such as X-tremeGENE siRNA Transfection reagent, can provide a convenient, reliable, and efficient vehicle for delivering siRNAs into animal cells, enabling the study of cellular and functional consequences of gene knockdown. This innovative reagent forms a complex with siRNA, as well as with mixtures of siRNA and plasmid DNA (cotransfection), and efficiently delivers the nucleic acids into animal cells to induce gene silencing. Transfection is achieved in just a few steps: mix and incubate diluted transfection reagent with diluted siRNA, then add this complex to cells. Because X-tremeGENE siRNA Transfection Reagent functions exceptionally well in the presence or absence of serum and demonstrates low cytotoxicity, it can be used without media changes. The product is animal-component free. Contents Solution filtered through 0.2 μm pore size membrane, supplied in polypropylene tubes. |
| Legal Information: | X-tremeGENE is a trademark of Roche |
| Other Notes: | For life science research only. Not for use in diagnostic procedures. |
| Quality: | Activity assay: X-tremeGENE siRNA Transfection Reagent (1 - 2.5 μl) is combined with siRNA (0.1 - 0.35 μg) that is specific for the HPRT housekeeping gene. The mixture is used to transfect HEK-293 cells (in a monolayer, 30 - 50% confluent) in the presence of 10% fetal bovine serum (FBS). Following transfection, the decrease of HPRT mRNA cells is determined with the LightCycler® Real-Time PCR System. A knockdown efficiency of 70 - 95% is typically observed when mRNA is measured. Cytotoxicity analysis: HEK-293 cells are exposed to siRNA/X-tremeGENE siRNA Reagent complexes for 72 hours in the presence of serum, without a media change. Cytotoxicity is then tested by analyzing the cells with the WST-1 Cell Proliferation Reagent (Roche). |
| RIDADR | NONH for all modes of transport |
| WGK Germany | nwg |
| Flash Point(F) | does not flash |
| Flash Point(C) | does not flash |
| Storage Temp. | 2-8°C |
| UNSPSC | 41106502 |