General siRNA FAQs

What is siRNA?

Small interfering RNA (siRNA) is a class of small double-stranded RNA molecules that silence gene expression by inducing the RNA-induced silencing complex (RISC) to cleave messenger RNA (mRNA)

What is the average molecular weight of my siRNA?

The average MW of a 21 bp duplex for siRNA is ~13,300 g/mol. The actual MW is listed on the Quality Assurance Document in g/mol.

What are the recommended storage conditions for siRNA duplexes?

Dried siRNAs are shipped at ambient temperature. Upon receipt, prior to reconstitution, store at –20 °C. For extended storage of reconstituted product, freeze at –20 °C in working aliquots. Repeated freezing and thawing or storage in “frost-free” freezers is not recommended.

How long are siRNAs stable?

Our R&D performed an accelerated stability study on several siRNA duplexes. They found that dry siRNA duplexes kept at -20 °C for three years are stable. Dry siRNA duplexes kept at room temperature only showed signs of minor degradation after two years. Our study did see some sequence-specific stability differences--only some sequences showed any signs of degradation at the two year mark. This is only representative of duplexed siRNA.

How many times can I freeze and thaw reconstituted siRNAs?

Repeated freezing and thawing or storage in “frost-free” freezers is not recommended. siRNA may be stored for up to six months at - 20 deg C in a non-frost-free freezer, we recommend that the solution is not freeze/thawed more than 3-5 times. In the lyophilized form the duplexes stable for much longer at - 20 deg C.

What level of purification do I need for my siRNAs?

In the vast majority of cases desalt (DST) siRNA duplexes will more than suffice to meet your needs. The process of purification basically enriches oligonucleotides for the full-length sequences. Our siRNAs are duplexed together from single-stranded synthetic RNA molecules that are individually "desalted" to remove by-products of the synthesis reactions, quality controlled, and quantified. The duplexing of the separate strands self-selects for the full-length oligonucleotides, as this process relies upon a perfect match between the self-complementary strands. Depending on manufacturing site, the duplex itself may then be separately run on a non-denaturing PAGE gel to check its overall integrity. Contact technical service if you have additional questions.

Do you offer GMP siRNA?

Not at this time

How do we anneal siRNAs to form a duplex?

We use the Tuschl protocol. It has been validated internally. For annealing, we incubate 20 µM single-stranded 21-nt RNAs in annealing buffer (100 mM potassium acetate, 30 mM HEPES-KOH at pH 7.4, 2 mM magnesium acetate) for 1 min at 90 °C, followed by 1 hour at 37 °C. The solution can be stored frozen at -20 °C and freeze-thawed many times. If a higher concentration of siRNA has been requested, we will use more than 20 µM of each strand.

Why does my calculated amount of siRNA in solution differ from that on the Quality Assurance Document?

There are multiple reasons why this may be the case: 1. The sample may not be homogeneously mixed. 2. Differences in instrumentation for quantifying siRNA may lead to differences in apparent values. Dual beam UV-VIS spectrophotometers are recommended. 3. The sample may be too concentrated. Absorbance values are most accurate between 0.15 and 0.6 and within the linear range of a standard curve. 4. The sample may be too diluted. Measurements with dilutions of small volumes (1-1.5 μL) are more susceptible to variation.

We received the siRNAs 2 weeks ago, and it is sitting on my benchtop. Are my siRNAs still ok?

Yes. Samples are shipped as dried pellets and are stable at room temperature for 2-4 weeks. Upon receipt, we recommend to store siRNAs at -20ºC or -70ºC to -80ºC.