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siRNA and miRNA Transfection: INTERFERin®

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  • Great silencing using as little as 1 nM siRNA
  • Suitable for miRNA transfection
  • Over 90% gene silencing in a wide variety of cells
  • Easy standard protocol
  • Gentle mode of action for more robust data and excellent cell viability
  • Compatible with serum and antibiotics

INTERFERin® provides more than 90% silencing efficiency at 1 nM siRNA in a wide variety of cells; hence avoiding off-target effects.

For High Throughput Screening Applications, Polyplus-transfection developed a dedicated reagent: INTERFERin®-HTS.

More information:

Expand All Tabs
Catalog Number Amount of reagent
409-01 0.1 ml
409-10 1 ml
409-50 5 x 1 ml
1 ml of INTERFERin® is sufficient to perform 500-1000 transfections in 24-well plates. Bulk sizes available, please contact us.

 

Several publications show that using low siRNA concentrations avoids off-target effects observed when transfecting siRNA at higher concentrations1,2. Ensure more reliable data by minimizing these unwanted non specific side-effects by using as little siRNA as possible. This is why INTERFERin® has been specifically designed to provide high silencing efficiency using low siRNA amount.
INTERFERin®-mediated delivery of 1 nM of a specific siRNA shows selective and highly efficient knockdown of gene expression, while a competitor (L2K) needs at least 10 nM siRNA to reach 50% silencing efficiency (Fig 1).

Fig 1. INTERFERin® only requires 1 nM siRNA for efficient gene silencing. 3LL cells stably expressing firefly luciferase were transfected with an anti-Luc siRNA using INTERFERin® or competitor L2K according to the manufacturer’s recommendation. Luciferase expression was measured after 48 h using a conventional assay. No inhibition was observed with control siRNA.

Amazingly, 50% silencing is still achieved at 10 pM siRNA (Fig. 2).

INTERFERin-fig1 Fig 2. INTERFERin® is highly efficient even at extremely low siRNA concentration. A549-GL3Luc cells were transfected in the presence of serum with decreasing concentrations of an anti-Luc siRNA using INTERFERin®. Luciferase expression was measured after 48 h using a conventional assay. No inhibition was observed with control siRNA.

Transfection of 1 nM siRNA targeting endogenous lamin A/C with INTERFERin® drastically reduces lamin gene expression to barely detectable level (Fig. 3).

INTERFERin-fig2 Fig 3. Transfection of 1 nM siRNA with INTERFERin® results in efficient and specific gene silencing.  CaSki cells were transfected with 1 nM lamin A/C siRNA using INTERFERin®. After 48 h, lamin A/C silencing efficiency was determined by immunofluorescence microscopy.

 

Transfection of microRNA (miRNA) oligonucleotides is increasingly being used to analyze biological effects of specific miRNAs on cell function. INTERFERin® is the reagent of choice for delivering miRNA, miRNA mimics or pre-miRNAs.

INTERFERin® is the latest generation siRNA & miRNA transfection reagent, especially designed for high transfection efficiency in a wide variety of cells, resulting in high gene silencing or stimulation of gene expression. Indeed, some miRNA are also known to induce gene expression by association with the promoter of the gene of interest. For example, INTERFERin®-mediated delivery of miR-373 leads to a 3 fold increase in E-Cadherin expression (Fig 4).

Figure 1

Fig 4. Transfection of miR-373 with INTERFERin® enhances E-cadherin expression. PC3 cells were seeded in a 6-well plate and transfected with miR-373 (25 nM) using 8 µl per well of INTERFERin®. 72h after transfection, E-cadherin mRNA expression level was determined by RT-qPCR. The assay was normalized with the HPRT-1 gene expression.

The cationic components of INTERFERin® require very little oligonucleotide amount, resulting in a gentle transfection process. In addition, the easy and robust protocol ensures reproducible results. miRNA can be transfected with small amounts of INTERFERin® resulting in high gene silencing or stimulation of gene expression. Whatever the concentration of INTERFERin® used, the effect stays approximately the same making INTERFERin® an economical reagent. For example, the fold change using different volume of INTERFERin® is always between 1,5 and 1,8 (Fig 5).

Figure 2 v2

Fig 5. INTERFERin® can be used at a very small amount of reagent. PC3 cells were seeded in a 6-well plate and transfected with miR-373 (10 nM) using 4, 6 or 8 µl per well of INTERFERin®. 72h post-transfection, E-cadherin expression was analyzed by Western Blot. The assay was normalized using β-actin expression. The quantification was expressed by taking the control situation as reference.

High transfection efficiency associated with high cell viability makes INTERFERin® the reagent of choice for generating relevant data for scientific publications. Indeed, the number of publications using INTERFERin® for miRNA and miRNA related molecules has been exponentially growing since its launch (Fig 6).

Evolution of publication number - Copie

Fig 6. INTERFERin® is involved in an increasing number of publication about miRNA transfection (Google Scholar).

 

For adherent cell lines or primary cells, 1 nM siRNA is sufficient to obtain more than 90 % gene silencing. For suspension cell lines, 80% silencing can still be reached by INTERFERin® using 5 nM siRNA (Table 1).

Table 1. Successfully transfected cell lines and silencing efficiencies obtained with INTERFERin®.

 

When it comes to cell viability, INTERFERin® outperforms other transfection reagents. 48 h after transfection with 1 nM siRNA, cells transfected with INTERFERin® appear healthy, while toxicity is clearly observed with reagent S (Fig. 7).

INTERFERin-fig4 Fig 7. INTERFERin® is extremely gentle on cells. Comparison of cell morphology 48 h after siRNA transfection using INTERFERin® or competitor reagents. A549-GL3Luc cells were transfected in the presence of serum with 1 nM Luciferase siRNA using INTERFERin® or competitors S or H according to the manufacturer’s protocol

 

INTERFERin® is ready to use and the protocol is straightforward. Start with 1 nM siRNA as a starting concentration for silencing of most genes in most cell types (Fig. 8).
INTERFERin® is compatible with both serum and antibiotics, hence forget time consuming medium changes and washes; INTERFERin® can be left on the cells without any adverse effects.

Fig 8. INTERFERin® standard protocolfor 24-well plate INTERFERin protocol 2012 2

Take a look at our Expert Tips in Genetic Engineering and Biotechnology News.

 

1.    Birmingham, A. et al., (2006) Nature Methods, Vol.3, No3, 199-204
2.    Caffrey, D. et al., (2011) PLoS One. 2011;6(7):e21503. Epub 2011 Jul 5.