RNA Oligonucleotides

Calculating this conversion requires you to know the extinction coefficient for your sequence. With this, you can easily convert from your nanomole amount to ODs. To determine the extinction coefficient, you can analyze your sequence using IDT's free OligoAnalyzer™ Tool. Results from this analysis will also provide you with nmol/OD₂₆₀ and µg/OD₂₆₀ values.

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Yes. If you can provide the aptamer sequence(s), you can order directly from the IDT website.

To order online click on Custom DNA oligos, or if your sequence contains RNA bases, Custom RNA oligos.

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Purification removes truncated products and other synthesis impurities. Our experience has shown that purifying an oligonucleotide that will be used in demanding applications saves both time and money in the long run. We recommend considering additional purification for any oligonucleotide that will be used for an application other than routine PCR, qPCR, or Sanger DNA sequencing.

As a general rule, IDT also recommends that any oligonucleotide longer than 40 bases should receive further purification. For questions, please contact us.

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Additional purification for oligonucleotides incorporating randomized bases can result in a bias in the degeneracy of the randomized bases.

Oligos with degenerate/randomized bases do not migrate in a tight distribution. Purification by either method (PAGE or HPLC) may purify out some of the randomized sequences. This results in the reduction of sequence diversity.

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Every IDT custom oligo is synthesized to order through a series of tightly controlled steps, including the coupling of the individual bases, cleaving the oligo from the solid support, desalting, and, if requested, purification. Each of these steps will result in a loss of final yield, which varies with each synthesis.

Due to this variation, IDT oligos are ordered according to the amount of starting material, also referred to as the scale. All custom oligos are assigned a minimum yield guarantee, but lot-to-lot variability in the final delivered quantity should be expected even when reordering the same sequence. For further questions about this subject, contact us.

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Customers who are new to working with large amounts of material are sometimes surprised when they see a yellow or brown discoloration to their oligos. If you have experienced this, do not worry. Color variation is normal in custom synthesis, especially as concentration increases. Importantly, IDT does not expect this to affect oligo function in our customers' experimental applications.

The image above depicts normal color variation as seen in oligo pools which are formulated to the same concentration (200 nmol oligos in 15 mL solution). In general, modified oligos at larger scales are more likely to display the variation seen above, while small-scale unmodified oligos (e.g., 25 nmol) tend to remain colorless in solution.

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IDT oligos have various estimated ship dates depending on product type and composition. Oligos added to the shopping cart will show an estimated ship date for each line item.

Once the order is placed, this estimated ship date can be found on the order confirmation email as well as in the order history of your web account. You can find the order history at idtdna.com/site/orderstatus/orderstatus.

Select the order number you wish to review, and the status and estimated ship date of each line item will be shown.

IDT will email you a confirmation when your order has been shipped. This shipping confirmation will include the tracking number of the package, which you can use to monitor the status of your shipment. If you have additional questions about your order, please contact us.

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To determine the relative T_m of primers with non-complementary overhangs, only the complementary region should be taken into account.

You can obtain the T_m using the free, online OligoAnalyzer™ Tool.

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The T_m value reported on our spec sheets uses a 25 µM oligonucleotide and a 50 mM salt concentration. It does not take into account dNTP or Mg²⁺  concentrations.

To get an accurate T_m for your specific application, all of these concentrations should be entered and adjusted to match the reaction conditions you plan to use.

The free IDT OligoAnalyzer™ calculates T_m for the concentrations you input.

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IDT uses a proprietary desalting technique that removes some truncation products and small organic contaminants from the synthesized oligonucleotide preparation. Removal of n-1mers produced during synthesis requires additional PAGE or HPLC purification.

PAGE is recommended for unmodified oligos >80−100 bases, while HPLC is the preferred method for oligos modified with either fluorescent dyes or attachment chemistries.

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RNA is inherently less stable than DNA due to its chemical structure. Additionally, RNases are more prevalent in standard laboratory conditions than DNases.

As even the slightest exposure to RNase can impact RNA stability, IDT has not performed rigorous long term stability studies for RNA.

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The sodium salt exchange we perform post-HPLC purification is a sodium acetate precipitation. It helps remove any remaining TEAA in the oligo preparation after HPLC purification.

It is important to make sure oligos used in in vivo assays are TEAA-free because it can be toxic to cells.

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The shelf life of an oligo is dependent on the temperature at which the oligo is stored and how the oligo is resuspended. Temperature is the more important of the 2 variables. Generally, oligos should be stored at –20°C. At this temperature an oligo has a minimum shelf life of 2 years, whether it is stored dry/lyophilized, in TE buffer, or in (non-DEPC treated) water.

Please see our DECODED™ article, Storing oligos: 7 things you should know, for data on oligonucleotide storage and a more thorough explanation.

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Oligo synthesis is accomplished through a series of steps, including coupling of individual bases, cleaving the oligo from the solid support, desalting, and if requested, purification of the oligo by HPLC or PAGE. No chemical reaction occurs with 100% efficiency, and, thus, each of these steps will incur a loss of final yield, which varies from specific sequence synthesis to synthesis.

Due to this variation, IDT custom oligos are ordered according to the amount of starting material used for the synthesis, referred to as the scale. While we cannot predict the actual final yield, we do guarantee a specific minimum yield for each oligo based on its sequence, starting scale, and the typical yield obtained under those specified conditions.

If you would like to know the minimum guaranteed yield for a specific oligo, simply add it to your Shopping Cart. The Shopping Cart provides you with the guaranteed minimum yield. If it is insufficient or excessive for your needs, simply edit the scale.

Please contact us if you have any questions about the yield you have received.

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For most applications, it is best to purify PCR products by gel electrophoresis.

This simple method not only purifies the final PCR product but can be a valuable troubleshooting tool as well since nonspecific PCR products, primer-dimer products, negative amplification, and other elements can easily be identified by gel analysis.

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The heterocyclic ring structures in DNA and RNA absorb light with a maximum absorbance near 260 nanometers (nm). An OD₂₆₀, or optical density 260, is defined as the amount of light at a 260 nm wavelength which will be absorbed by an oligo resuspended in 1 mL water and the concentration is read in a 1 cm quartz cuvette.

This method of measurement is considered the most accurate means of assessing the amount of oligonucleotide present following synthesis. The relationship between measured OD₂₆₀, molar extinction coefficient (ε260), and oligonucleotide concentration is given as: OD₂₆₀ = ε260 x concentration.

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IDT uses our own proprietary HPLC method to purify oligos in-house. A published set of conditions for an RP-HPLC protocol is:

• Hamilton PRP-1 Reverse Phase column 250 x 10 mm
• Buffer A = 100% ACN (acetonitrile)
• Buffer B = ACN/0.1 M TEAA pH 7.3
• Run at 4 mL/min over 30 min from 95–60% A
• Monitor at 260/297 nm

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Ideal amplicon length/size depends on many variables and design preferences.

For standard PCR, scientists generally design amplicons to be between 200–1000 bp.

For quantitative PCR, standard amplicons range from 75–150 bp. It is unlikely that an amplicon will be too short. However, amplicons of >1000 bp may need extra time to be completed during the extension step.

The polymerization rate of the enzyme used in the reaction needs to be sufficient to copy the amplicon in its entirety, so for long amplicons, you may need to increase the extension cycle time.

The general rule of thumb is 1 min/kb of amplicon. You will also need to optimize reaction component concentrations based on the amplicon size.

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