Tm is shorthand for melting temperature. The melting temperature of a nucleic acid duplex is the temperature at which half of the strands are in duplex form, and the other half are single stranded. This is determined by taking advantage of a spectral phenomenon called hypochromicity, which is the reduction in UV absorbance that is observed when nucleic acid bases form a duplex. The act of hydrogen pairing with the opposing strand and the change in electron patterns results in the observed reduction in the spectral reading at 260 nm. By cooling a sample containing complimentary strands in a cuvette until the strands are fully hybridized (absorbance is stable), then heating the sample slowly and recording the absorbance during the process, one obtains a sigmoidal shaped plot when temperature is plotted against absorbance. The transition temperature at the inflection point is the Tm.
What is the relative order of stability of the basic duplex combinations?
2’OMe:RNA > RNA:RNA > DNA:RNA > DNA:DNA. For further detail, refer to our article Thermostability of Modified Oligonucleotide Duplexes.
How do I hybridize my oligonucleotide?
Resuspend the two complimentary strands in a neutral pH buffer. Measure the absorption of the solution using a spectrophotometer and calculate the concentration in moles using the provided extinction coefficient. Mix the two oligonucleotides together in equal molar ratios. Hybridization can be confirmed by running 3 samples on a gel under native conditions. Sample 1: single stranded oligonucleotide 1; Sample 2: single stranded oligonucleotide 2; Sample 3: hybridized oligonucleotide 1 and oligonucleotide 2. If hybridization was successful, the hybridized oligonucleotide Sample 3 lane will run slower compared to the single stranded Samples 1 and 2. If a little of one of the strands is observed in the duplex lane, titrate with the other strand until only duplex is observed. In some cases, it will be necessary to chill the gel to ensure duplex stability.
What oligonucleotide QC analyses do you provide?
We confirm quality of each oligonucleotide, regardless of level of purification, by PAGE analysis and our in house mass spectral analysis. Please contact us to review data from a previous order. For complex oligonucleotides that require mass spectral analysis not available at TriLink, please request mass spec at time of quote or order. An additional fee may be required. For mid-scale (50 mg or greater) syntheses, we also offer HPLC analysis at no additional cost.
We offer a variety of analyses including HPLC (AX, RP or Ion Pairing), Gel Densitometry, Conductivity and Enzymatic Digest. Please request any additional analysis at time of quote or order as an additional fee may be required.
We will often obtain data during the process that we do not routinely provide the customer, such as the uv/vis absorption spectrum of dye labeled oligonucleotides. If you would like to see a particular piece of data, please contact us, and we will do our best to accommodate you. If you would like to see this data routinely, we will add it to the specifications for your program and include any additional fees as required.
Do you supply a certificate of analysis, and if so, what is on it?
We supply a certificate of analysis with every oligonucleotide shipped. The certificate of analysis contains the sequence and modification information, yield, purity with assay specifics, a copy of the actual analytical data, usually PAGE, and certain physical data, such as the molecular weight and calculated extinction coefficient.
Can I purchase the phosphoramidites that you list as modifications for your custom oligonucleotide products?
We prepare many of our linkers and other 5’ and 3’ modifiers in-house. They can now be purchased directly from Glen Research. The modified base phosphoramidites used in our oligo synthesis lab are not made in-house. They can also be purchased through Glen Research.
Does TriLink offer primers?
Yes we do. Although we do not have high throughput capabilities for orders of thousands of primers, we can handle most of your needs, and give you the high quality you have come to expect from TriLink.
What is the turnaround time for receiving my oligonucleotide once I place my order?
The amount of time depends on the type of oligonucleotide you order. Small-scale, unmodified oligonucleotides can take as little as three days. If purification is requested, the oligonucleotides will take slightly longer to be shipped. A midscale order (50 mg – 10 g), will take approximately four weeks. Highly modified or conjugated oligonucleotides can also take longer. We always ship your oligonucleotides out as quickly as possible, but we refuse to sacrifice quality. Upon placing the order you will receive a confirmation email that typically includes an estimated shipping date. We will update you as soon as possible if there are any changes to the estimated shipping date.
What is the difference between ordering an oligonucleotide at a scale amount versus in milligrams?
Ordering by a scale amount means you are requesting a starting scale synthesis. The final yield will not equal the starting amount, since there are synthetic, purification and processing losses that differ from compound to compound and often from batch to batch of the same material. Because of this, the price is calculated based on the pre-determined cost of material and labor for a synthesis at that particular scale. Our small-scale orders include all syntheses up to the 15 µmole scale. When you place an order in milligrams or grams you are requesting an amount of final product. We have set prices starting at 50 mgs for large-scale orders. If the oligonucleotide has any modifications, then a custom quotation will be necessary. You may request a specific final yield for the smaller scale syntheses, such as 1 μmole, however please keep in mind that it may require a 5 µmole starting scale or more to guarantee that yield. When you specify the final yield, we must begin with a large enough scale to ensure the yield is sufficient. Prior experience with your particular compound will help us determine a fair price.
Does my product pose a ship hazard?
All of our stocked items are non-biological, non-hazardous research reagents and do not pose a ship hazard. For custom synthesized compounds please contact us to determine if there is any issue or concern.
How do I obtain an MSDS for my product?
For our stocked items the MSDS is available online in the product specific page. For custom synthesized compounds please have the lot number available and contact us. We will be more than happy to supply one for you.
Can you provide a Proforma Invoice?
Yes. Please contact us to request a proforma invoice.
Is payment due when I place my order?
No, you will not be billed/charged until your order is shipped. We accept payment by purchase order, wire transfer and credit card (VISA, MasterCard and American Express). Terms are net 30. Please contact us for more information.
Why is there a difference in pricing for molecular beacons depending on whether I have a license or not?
Our license to the technology requires that we pay a royalty for molecular beacons sold to institutions that do not have a license from The Public Health Research Institute of New York (PHRI). If you have a license, we do not have to pay a royalty to PHRI on your beacon. When you place your first order for a molecular beacon, we will ask if you have a license from PHRI.
How do you release products for shipping?
The batch record for your product is first reviewed and signed by the production manager in charge of manufacturing the product. All products are reviewed by a quality assurance specialist who signs the certificate of analysis verifying that all appropriate quality systems were followed and that all specifications were met prior to shipment.
Is there a list of restricted countries to which you cannot ship?
| Shipping restrictions vary for specific countries and items. We recommend using a forwarding service for all shipments whose destination is not located in North America, Europe or Australia. This is mandatory if the shipment requires dry ice. We recommend using Biocair forwarding service; more information is available at https://www.biocair.com/. Please inquire about your particular location. |
How much does shipping cost?
Shipping costs vary depending on your location, whether or not dry ice is required, as well as the size of the dry ice box. All products delivered in solution will be shipped on dry ice. We ship via FedEx Priority Overnight or FedEx International Priority. Within the United States, standard shipping may range from $35-$350. International shipments have a greater range due to distance. If your order requires dry ice or insurance, additional charges will apply. Please inquire for pricing specific to your location and order.
Why is your turn around time longer than I expected?
The simple answer is that it takes longer to make a high quality, modified oligonucleotides reproducibly. TriLink was formed in 1996 largely in response to the poor quality of modified oligonucleotides available from many vendors. We developed very methodical protocols geared to the manufacture of highly modified products. Our protocols require a great deal of attention to detail, as much of the work must be done manually to ensure success. Additionally, our purification methods are very time consuming, and we include 1-2 days for the analysis of products prior to shipping.
How will my order be shipped?
We ship via FedEx Priority which guarantees receipt by 10:30 am the following business day to most locations within the US. Our oligonucleotides are packaged in microcentrifuge tubes as lyophilized solids. Oligos are most stable lyophilized; however, we can ship them in solution in water or buffer upon request (an additional charge may apply). Unless requested, all oligos are shipped at room temperature. They are stable at room temperature, but should be stored at –20°C upon arrival. Compounds in solution, with a dimethoxytrityl group must be shipped on dry ice to ensure stability of the trityl group. Please note: dye labeled oligos are light sensitive. All oligos are shipped in an inner reflective Mylar envelope to shield them from light. This envelope can be used for storage.
Can I speed up the delivery of my order?
Orders are prepared on a “first come, first served” principal. We do our best to accommodate your wishes by streamlining the process, however, our processes are generally firmly set, by both instrumentation and the training of the staff. Be aware that changing protocols may compromise the quality of the product. Most nights are already used for longer steps, such as deprotections, or dry-downs of larger samples. We can often schedule overtime if your requirements are truly urgent and you are willing to pay the added costs. This can save no more than a couple of days, since delivery is usually within 2 weeks. However, please discuss this option with us if you are interested.
Do you give academic/university discounts?
No, not automatically. Universities are eligible for bulk and volume discounts as single organizations. If you have a number of buyers in your university, talk to your purchasing agent about obtaining an Annual Purchasing Agreement for the whole university.
How can I get a discount?
You can get a discount by signing an Annual Purchase Agreement (APA). Based on your forecasted order volume, you will be eligible for a discount for the following twelve months. Some products are already heavily discounted and are excluded from the Annual Purchase Agreement, and custom compounds are quoted on a case-by-case basis at the best price currently possible. Bulk discounts are also available for many products. Call or email for details.
How do I set up an account for placing orders?
We will set up your account at the time of your initial order. You will need to provide a PO number or a credit card number and submit a credit application. Our accounting department will contact yours to obtain the remaining information required to establish credit. We retain the right to deny credit to any customer at any time.
Can I make changes to my order once it has been placed?
NTPs are shipped the following business day, unless otherwise stated. Changes or additions may be made to your order until the time of shipping, which is 3pm PST. For oligonucleotide or custom compound orders, if we can pull the order out of production before synthesis is started, we can accommodate your request without incurring an extra fee. However, once synthesis of the product has begun, changes to the sequence or scale are no longer possible unless costs of the current synthesis are met. Please keep in mind that you may be responsible for the cost of any specialized reagents that were purchased to synthesize your compound, plus a 10% surcharge. PAGE or HPLC purification may be added anytime before an oligonucleotide is shipped.
Does TriLink have distributors?
Yes, click here for a list of distributors by region.
How do I place an order for TriLink products?
1) Use our secure online ordering system (Link changed)
2) Email us
Do you offer small-scale oligonucleotides under GMP conditions?
Yes. We offer full GMP quality material at all scales, as both diagnostic grade and pharmaceutical grade.
What kind of nucleotides can you prepare under GMP conditions?
We can prepare any kind of nucleotide in our GMP facility. We first develop, if necessary, the process in our R&D lab, then validate it in our GMP facility with our equipment and reagents. Nearly all TriLink products are diagnostic grade GMP and manufactured in compliance with ISO 9001.
Do you offer pharmaceutical grade products?
We are designing a new facility that will be suitable for the manufacturing of pharmaceutical grade materials by early 2014.
Which level of GMP do I need?
This is a decision for your regulatory department. However, we can offer advice based on your specific circumstances if you are unsure. Often, you do not need as stringent a manufacturing system as you may first believe.
Can I get research grade material that is well documented, but not as expensive as GMP grade material?
Yes! We find the costs of managing separate documentation systems (one for research grade material and one for GMP grade material) are more than the cost savings associated with the reduced documentation required for research grade materials. Therefore, at TriLink there’s no such thing as research grade material. All compounds are manufactured under a GMP system at comparable prices to research grade material from our competitors.
Is TriLink ISO certified?
Yes. We are ISO 9001 certified.
What do GMP, ISO and QSR mean, and how do they differ?
GMP stands for Good Manufacturing Practices, and refers to a system of manufacturing that guarantees reproducibility of product quality to set specifications. cGMP is simply Current Good Manufacturing Practices and refers to compliance with current regulations. It can be considered redundant since to be GMP compliant you must comply with current GMP regulations anyways. The requirements differ depending on what type of product is being manufactured and whether it is for pharmaceutical or diagnostic purposes. ISO stands for International Organization for Standardization, which offers a standard for operating a firm from management through manufacturing. It is more encompassing than GMP. QSR stands for Quality Systems Regulation, which are GMP standards described by the FDA for the manufacture of products for the diagnostic industry. The ISO and QSR systems each describe specific GMP standards. The ISO system pays more attention to the management of the firm and places a number of reporting loops in the firm to ensure attention to issues. The QSR system is more focused on the manufacturing systems, and the validation of those systems. Although neither standard is required to maintain GMP facilities, it is essential that a firm satisfies the requirements of the client’s quality system. Generally, this means conforming to ISO or QSR standards. Every product made at TriLink is manufactured under GMP. We are compliant with ISO 9001.
What scale of products do you offer?
We offer up to 10 grams per lot of most oligonucleotides and 100 grams per lot of modified nucleotides. Please inquire about our current capabilities.
Where are you located?
We are located in San Diego, CA, in the Sorrento Mesa area just east of I-805 and just north of Miramar Marine Air Station. Our address is 9955 Mesa Rim Rd. San Diego, CA, 92121. Map It
What are TriLink’s hours?
We are open from 7:00 AM to 5:00 PM PST (10:00 AM to 8:00 PM EST).
Will I receive a discount on multiple modified base insertions in the same oligonucleotide?
Yes, if you order an oligonucleotide with multiple couplings of the same modified base your price will be adjusted accordingly after your order is placed. You will recieve a 25% discount on the second coupling and a 50% discount on all additional couplings of a particular base within a single oligonucleotide.
Oops! How do I go back and add a modification in the middle of my sequence – it keeps inserting at the end?
The Internal Modifications click menu works by inserting the modification at the current end of the oligonucleotide you are building. If you need to go back and insert an internal modification after you have filled in your entire sequence, you will either need to:
1. Delete back to the position of the modification, select it in the menu and fill in your remaining sequence, or
2. Select the modification, then carefully cut and paste the modification into the correct position.
Please always be sure to double check your sequence before placing your order.
How do I order a CleanAmp™ Primer?
CleanAmp™ Primers can be ordered here.
How do I insert a dye or conjugate on an internal linker?
Several modifications are available in the amidite form and can be added to your sequence simply by clicking on them. These modifications are marked “Amidite.” For all other internal dyes and conjugates you will need to choose the appropriate selective placement linker. To add an internal dye or conjugate click the linker of choice, followed by the dye or conjugate of choice from the internal modifications menu. The modification will appear at the end of your current sequence. Continue adding bases as needed. To view all available selective placement linkers, dyes and other non-fluorescent conjugates, see our Custom Oligonucleotide Components menu.
Can I order an oligonucleotide with a modification I do not see in the list?
TriLink offers numerous modifications that do not have standard list pricing and therefore do not appear in OligoBuilder®. These include 2’ Fluoro RNA bases, symmetrical branchers, cis-syn thymidine dimers, trimers and many more. Please email sales@trilinkbiotech.com for a quotation on any oligonucleotide modifications you do not see in OligoBuilder®. Be sure to include the sequence, backbone, modifications, starting synthesis scale or final yield and purification requirements.
How much material should I expect from my starting synthesis scale?
Click here, then select your backbone to view expected yields for each.
How do I know which purification method to choose?
Click here to learn about each purification option. If you are not sure which purification is most appropriate for your construct and application, select “Best Method” and our tech support team will assist you.
How do I know which backbone option to choose?
Click here to view backbone details.
Is there a length limitation to the aptamer sequence I can have chemically synthesized?
Yes, due to secondary structure formation chemical synthesis becomes challenging around 70 bases. However, under TriLink’s RNA transcription service much longer aptamers can be made. Learn More>
I noticed you offer libraries with varying lengths of random regions, which one should I use?
Libraries with longer random regions have more unique sequence motifs than do libraries with shorter random regions. However, not all possible unique sequences can be represented in each selection. For example, a library with a 30 nucleotide random region has the potential for 1.1 x 1018 unique sequences and a library with a 20 nucleotide random region has 1.1 x 1012 unique sequences. Typical selections start with 1015 copies of library. For a library with 20mer random region, approximately 1,000 copies of each unique sequence are present, while for the corresponding library with a 30mer random region, only one out of every 1,000 unique sequences is represented.
In contrast, libraries with shorter random regions will give you a better representation of all possible sequences but are inherently less complex than a library with a longer randomer region. However, once an aptamer is selected shorter aptamers are easier and less expensive to synthesize.
We offer libraries with different random region lengths, allowing the right balance of library complexity and library representation to be experimentally determined for your selection.
What is the ratio of TriLink’s wobble bases? Is this important?
TriLink’s random sites are optimized to produce as close to a 1:1:1:1, A:C:G:T base ratio in the resulting wobble site as possible. TriLink has done extensive research in the area of randomer oligonucleotides. In aptamers any bias can cause misrepresentation of the sequence space, which can limit the search for aptamers that bind the desired target with high specificity. The randomness of TriLink’s library manufacturing process was confirmed by NeoVentures and featured in the whitepaper Validation of Random Library for Aptamer Selection.
What is the effect of the fixed primer sequences flanking my random region?
Although one would assume the flanking fixed primer regions would play a major role in the resulting aptamer structure, it has been demonstrated, in a bioinformatics study performed in Dr. Andrew Ellington’s lab that these constant regions are only minimally involved in the structures of selected aptamers.
Despite these findings, a number of selections are performed using fixed regions which are shorter than traditional primer binding sites. We have designed our libraries to contain an Ndel site (CA/TATG) upstream of the random region and a Spel site (A/CTAGT) just downstream. This allows the use of restriction endonucleases and ligases in a workflow to minimize the role of the fixed sequence in the selection.
How do I make an RNA library from a DNA library?
An RNA Library can be made from the DNA Library for DNA Aptamer Selection with the optional T7-Promoter Forward Primer. The T7-Promoter Forward Primer is essentially the same as the forward selection primer, with the exception that it has the T7 promoter sequence on the 5′ end. Use the T7-Promoter Forward Primer in combination with the reverse selection primer in PCR to generate the corresponding DNA template which contains the T7 promoter sequence. This DNA template can be used to generate the RNA library by in vitro transcription using T7 RNA polymerase.
Do you have a nucleotide modification that I can use for PCR incorporation that prevents further DNA amplification but is also a reversible modification? -Pete
The only analog that has the properties you described is our CleanAmp™ dNTPs. Although the protecting group will not be stable during the elevated temperatures of PCR, certain enzymes such as the Therminator series of enzymes can incorporate this type of analogs. However, incorporation would need to be performed at low temperatures, such as at room temp. Then, a heat activation step can be used to remove the protecting group.
We are trying to introduce aptamers as sensitizers into electronic transducers. We are looking for an aptamer that can be immobilized onto a metal (not so important what metal), and selectively binds to a biomedically relevant analyte in aq. solution (saline, or buffer). Any suggestions? -Martin
The first step is to identify the aptamer for the analyte of choice using one of the many selection protocols available. We recommend that you use a random library with no more than 40 random bases. Longer ones will yield aptamers that will be more difficult and expensive to prepare synthetically, especially if the aptamer is heavily modified. You can review our stocked libraries to see if one may be used during your selection.
Once you have the sequence, it is fairly easy for us to modify your aptamer with a reactive moiety for conjugation to a metal. For instance, we can put sulfur on the aptamer for direct conjugation to a gold particle. Once you have identified the metal(gold is well established in the literature) we can help with the decision regarding chemistry.
Please contact us for more details.
We are currently searching for an RNA with a Poly (A) tail that can be used in our RNase test kit. Can you manufacture a pre-RNA with a 3’ poly(A) tail?
Thank you for your interest in our RNA transcription service. We believe our capabilities are only limited by customers imaginations. Please submit a quote request with the details of your project to initiate discussions with our Product Management Team.
Can you provide an RNA aptamer library to find riboswitches specific for my metabolites of interest? -Ruchi
We understand you are looking to perform an aptamer selection for a metabolite of interest. Once selected, you will put this into one of the intergenic regions. In the absence of metabolite, this region will be linear, while in the presence of metabolite, this region will fold into a complex structure. The presence of this structure will then block translation. Therefore, we believe you are asking if you can design your aptamer library to have a 5’- and 3’- end that is fixed to match the intergenic sequences, with a random number of nucleotides (N20, N30) in between. And the answer to this is yes! We would be happy to make a custom library for you. This construct can be priced and ordered through OligoBuilder.
Do I have to activate the amine group of the 5′ C6 Amino Modifier for conjugation or is the product delivered with the ready-to-conjugate amine group? – Grace
A 5′ amine modified oligonucleotide will be delivered ready to conjugate to a succinimidyl ester.
Please keep in mind that TriLink can perform the conjugation and purify the final product for you. We can conjugate most commercial succinimidyl esters or one you supply.
I usually order (DNP-TEG)-labeled probes and the QCs are a PAGE and mass Spec analysis. Can you provide a purity level with each order, that I could use as a reference? -Aurelie
We run mass spec and PAGE analysis on every oligo ordered as part of our quality system. The PAGE analysis allows qualitative assessment and mass spec confirms identity. An HPLC or gel densitometry is required for quantification of purity. HPLC and other analytics can be requested at the time of order. If you are not sure which analytical would be best for your requirement, just ask!
In regards to the DNP-TEG labeled oligonucleotides, we recommend requesting an AX-HPLC analysis.
How did you quantify the samples in your white paper “PCR incorporation of modified dNTPs: the substrate properties of biotinylated dNTPs?” -Josh
In this whitepaper, we quantified the amount of amplicon formed as a function of the amount of input biotinylated nucleotide by gel densitometry. In particular, the percent amplicon yield for a given percentage of biotinylated nucleotide was normalized to an reaction which contained only natural nucleotides.
While this approach can be used to identify optimal reaction conditions that will allow for biotinylated nucleotide incorporation in PCR without sacrificing amplicon yield, this does not measure the amount of biotin in the PCR product itself. Should direct quantification of the biotin content be of interest, enzymatic digestion of the isolated product is one way to do this. Eadie, et. al.and Andrus, et. al.describe this technique in their publications.
You state on your website that 6-FAM is less susceptible to photobleaching than FITC. Where can I find the reference for this? -Shane
A. Photobleaching of different fluorophores has been reviewed for use in a number of applications including quantitative microscopy and fluorescence photobleaching recovery (FPR). Dailey, et. al. reviewed studies of photobleaching in different systems, as well as different methods employed to reduce photobleaching.
While 6-FAM is less susceptible to photobleaching than FITC, newer dyes have been developed that are more stable. For example, Panchuk-Voloshina, et al. demonstrated the stability of Alexa Dyes over FAM and FITC.
What is the difference (especially in enzymatic incorporation and/or after incorporation as a template for PCR) between Biotin-AA-dCTP (N-5002) and N4-Biotin-OBEA-dCTP (N-5004)? I want to incorporate biotin in a primer extension reaction (Klenow), pull down with streptavidin, and then use the biotinylated template for PCR. – Jen
A. We have documented Biotin-AA-dCTP (N-5002) incorporating at a very high percent substitution for dCTP, up to 97%, in our white paper: PCR incorporation of modified dNTPs: the substrate properties of biotinylated dNTPs. We do not have analagous PCR data on the efficiency of the N4-Biotin-OBEA-dCTP (N-5004), but since the modification is to the N4 nitrogen, it may interfere with Watson-Crick base pairing. Therefore, while this dCTP analog can be incorporated in PCR, the percent substitution for dCTP may not be as high as for Biotin-AA-dCTP.
As you eluded to, different analogs will incorporate in PCR with varying efficiency and some optimization may be needed. We recommend performing some initial PCR experiments using natural:modified dNTPs in ratios such as 1:0, 3:1, 1:1, 1:3 and 0:1 to identify the best conditions for modified nucleotide incorporation with robust amplicon yield.
How versatile are CleanAmp™ dNTPs?
A. CleanAmp™ dNTPs can be used with a broad selection of DNA polymerases including Taq, Pfu, Pfu (exo-), Dynazyme™, Deep VentR™, Tth and Tfi. When using CleanAmp™ dNTPs with these polymerases, all were able to produce the desired amplicon. Learn More…
Do CleanAmp™ dNTPs form a modified amplicon?
A. No, CleanAmp™ dNTPs are reverted back to the corresponding standard dNTP prior to being incorporated into the amplicon. The 3’-hydroxyl modification blocks enzyme incorporation until it is removed during heat activation. After heat activation, the enzyme efficiently incorporates the corresponding standard dNTP.
What is a phosphoramidite?
A. A phosphoramidite is the actual building block we use to chemically synthesize oligonucleotides. It usually consists of a protected nucleoside with a 3’ phosphitylating reagent, where phosphorus is in the trivalent state and protected with a stable amine and ester. Learn More…
Can I buy linker reagents directly from TriLink?
Can I buy linker reagents directly from TriLink?
Yes, click here to view the reagents still sold through TriLink. All of our other DNA synthesis reagents are now sold through Glen Research.
Can TriLink label nucleotides with a DNA intercalating dye (e.g. Thiazole Orange)?
A. Yes, we are able to label nucleotides with a DNA intercalating dye. Using TriLink’s custom synthesis service, we can conjugate your dye of choice to dNTPs containing the appropriate linker. Read More…
Why does the price of trimer modified oligonucleotides vary?
Why does the price of trimer modified oligonucleotides vary?
Trimer modification requires a highly custom synthesis protocol in which multiple additional factors are taken into account. Each trimer mix job is individually analyzed in order to provide the most cost effective pricing in terms of synthesis cost and trimer amidite usage. Read More…
Why use CleanAmp™ dNTPs versus unmodified dNTPs?
A. CleanAmp™ dNTPs are able to decrease the formation of primer dimer products while increasing the specificity of the desired product better than unmodified dNTPs. Use of CleanAmp™ dNTPs also reduces data fluctuation, giving a more consistent PCR result.
Is it be possible to synthesize an RNA oligo of 60-90 nucleotides long?
Is it be possible to synthesize an RNA oligo of 60-90 nucleotides long? In addition, is it be possible to label some inner selected nucleotides as we desire? – Francisco Hernandez-Torres, University of Jaen
Yes, we can synthesize a 60-90mer RNA oligonucleotide. We have successfully made up to a 110mer unmodified RNA oligonucleotide and continue to try longer sequences as our customer’s request them. Yes, internal modifications can be incorporated. Read More…
How does Biotin-TEG RNA Oligonucleotide Help Dengue Virus Research?
DNA Methyltransferase (MTase) methylates the C-5 position of cytosines in DNA. This is a noteworthy family of proteins due to the implications of DNA methylations in the control of gene expression. MTases are a logical target for small molecule therapeutics for cancer and other diseases. Read Article
What is the longest oligonucleotide TriLink can synthesize?
What is the longest oligonucleotide TriLink can synthesize?
TriLink has successfully synthesized 220mer unmodified DNA and 110mer unmodified RNA. However, TriLink is willing to try longer sequences.
Post a comment below to discuss your needs.
What is the expiration date of my modified nucleotide?
What is the expiration date of my modified nucleotide?
All of our modified nucleotides are submitted to quality control annually to ensure that they still meet our specifications, therefore we do not assign expiration dates to our modified nucleotides other than our CleanAmpTM dNTPs. Most modified nucleotides are stable for several years when stored properly at -20oC. Read More…
Have a question about modified nucleotides? Post a comment below.
I was wondering if the following modified nucleotides can be used by Taq polymerase in PCR. 1-Thio-2′-Deoxyadenosine-5′-Triphosphate (N-8001) 1-Thio-2′-Deoxycytidine-5′-Triphosphate (N-8002) – Chun-Nan Chen, Single Cell Technology
1-Thio-2′-Deoxyadenosine-5′-Triphosphate (N-8001) and 1-Thio-2′-Deoxycytidine-5′-Triphosphate (N-8002) can be incorporated by Taq in PCR. However, not all analogs will incorporate with the same efficiency in PCR; some optimization may be needed. We recommend performing some initial PCR experiments using natural:modified dNTPs in ratios such as 1:0, 3:1, 1:1, 1:3 and 0:1 to identify the best conditions for modified nucleotide incorporation with robust amplicon yield.
What solution do you have for a DNA oligo with a fluorescent dye attached for a fluorescence polarization assay?
I need a DNA oligo with a fluorescent dye attached for an fluorescence polarization assay. The linker between DNA and dye should be as rigid as possible and the dye most likely a BODIPYdye, dansyl or fluorescein dye. Attachment either internal or at the 5′ of the oligo. What solution do you have for that? – Chris Gloeckner, Illumina
A shorter linker will provide the most rigidity between an oligo and a dye. Our shortest linker is the dT-C2 amino linker. The linker arm is attached to the Thymidine base and the dye would be conjugated onto the amino group post-synthetically. Another option would be to use our C7 internal amino linker. Read More…
Have a question about Oligonucleotides? Post a comment below.
Should I use the CleanAmp™ 7-deaza-dGTP Mix or just CleanAmp™ 7-deaza-dGTP alone?
Should I use the CleanAmp™ 7-deaza-dGTP Mix or just CleanAmp™ 7-deaza-dGTP alone?
For targets with <70% GC-content, addition of the CleanAmp™ 7-deaza-dGTP with standard dNTPs should amplify the target well. For targets with >70% GC-content, we recommend the CleanAmp™ 7-deaza-dGTP Mix for a robust amplification. Learn more about robust GC-rich target amplification in our white paper.
Have a question about CleanAmp™? Post a comment below.
What are the key benefits to new, improved CleanAmp™ dNTPs?
What are the key benefits to new, improved CleanAmp™ dNTPs?
Improved Amplicon Yield in Endpoint PCR
– Earlier Cq Values in Fast Cycling Real-Time PCR
Click here to learn more.
Have a question about CleanAmp™? Post a comment below.
Will CleanAmp™ dNTPs activate at 60°C?
Will CleanAmp™ dNTPs activate at 60°C?
We are currently testing the kinetics of the deprotection of CleanAmp™ dNTPs at lower temperatures such as 60°C. Both the temperature and the pH of the PCR reaction affect the deprotection rate of the CleanAmp™ dNTPs where increased temperature and acidic conditions accelerate deprotection. Read More…
Have a question about CleanAmp™? Post a comment below.
Is it possible to buy NTP’s (in particular ATP) that works like the CleanAmp™ nucleotides?
I just wondered if it is possible to buy NTP’s (in particular ATP) that works like the CleanAmp™ nucleotides? A preferable version would be activated by light. – Leif Larsen, Vipergen
The CleanAmp™ modification can be introduced onto a number of NTP analogs, including ATP. This would allow you to control activation via pH or heat. There are other approaches to introduce light activatable modifications onto ATP as well. Read More…
Have a question about NTPs? Post a comment below.
How do I reduce my thiol modified oligo?
How do I reduce my thiol modified oligo?
1. Make a solution of 60 mM TCEP. (18 mgs TCEP in 1 mL H2O)
2. Add 125 uL 0.1M NaHPO4, 0.15M NaCl, pH 7.26, vortex & make certain oligo is in solution.
3. Add 75 uL 60 mM TCEP to oligo solution.
4. Vortex.
5. Let sit for 2 hrs at room temperature.
6. Run size exclusion column, RP-HPLC.
Have a question? Post a comment below or email us at support@trilinkbiotech.com.
I don’t see the modified nucleoside triphosphate that I need, can you make it for me?
I don’t see the modified nucleoside triphosphate that I need, can you make it for me?
Yes, if you don’t see what you need on our website or in our catalog, please inquire about our custom chemistry services. In addition… Read More
Have a question about modified nucleoside triphosphates? Post a comment below.
I would like to know what condition (and aqueous solution) you recommend for suspension of the lyophilized oligos for in vitro use?
I would like to know what condition (and aqueous solution) you recommend for suspension of the lyophilized oligos for in vitro use? – Michael Dickinson
Prior to resuspending your oligo be sure to centrifuge it briefly to ensure all product is down in the bottom of the tube. We recommend bringing your oligo up in any aqueous solution with a neutral pH. If you use just water, be sure that it is nuclease free. Read More…
Have a question about oligonucleotides? Post a comment below.
How long does it take to activate CleanAmp™ dNTPs at 95°C and 60°C?
How long does it take to activate CleanAmp™ dNTPs at 95°C and 60°C? – Jia Wang
Thank you for your interest in TriLink. CleanAmp™ dNTPs have a t 1/2 of 40 minutes at 95°C. We are currently developing a new, improved version of CleanAmp™ dNTPs that have a t 1/2 of 7 minutes. We are also working towards an isothermal solution… Read More.
Have a question about CleanAmp™ dNTPs? Post a comment below.
I’d like to know the recommended condition to dissolve the synthesized TRIMER oligos? Is it better to use TE buffer, pH 8 or just DDI water? – Gene Xiang, Eureka Therapeutics, Inc.
We recommend resuspending oligonucleotides in aqueous, neutral solution. It would be best to resuspend your trimer oligonucleotide in DDI water. If you have any further questions, please do not hesitate to contact us.
Have a question about specialty oligonucleotides? Post a comment below.
An Antigene-based Oligonucleotide Therapeutic
Though the phenomenon of siRNA is now a key tool in drug development, it does have a fundamental disadvantage. Since siRNAs silence expression at the mRNA stage, it is presumed that siRNA drugs would need to be administered indefinitely. In a recent review article by N. Kolevzon and E. Yavin, the concept of a photoactivated TFO is proposed as a viable therapeutic option… Read More.
Have a question about therapeutic oligonucleotides? Post a comment below.
Do you have any CleanAmp products that were determined to be product “failures” during your development?
Do you have any CleanAmp products that were determined to be product “failures” during your development? I know you made products with a hot start at high temperature a goal but do you have any chemistries that activated at ~56oC and were never released as viable product? Customized chemistries are of interest for ongoing development in my organization with an activation temp ~56oC. Thanks, – Dan Shaffer, Envirologix
Thank you for your interest in our CleanAmp™ Products. We did, as you thought, try several protecting groups before landing on the current CleanAmp™ chemistry … Read More
Have a question about CleanAmp™ Products? Post a comment below.
Improving GC-rich Target Amplification
We are proud to introduce the latest addition to the CleanAmp™ Product Line, CleanAmp™ 7-deaza-dGTP for GC-rich target amplification. When DNA targets high in GC content are amplified, PCR product formation can often be compromised by inadequate strand separation and the propensity for complex secondary structure formation. The inability of the DNA polymerase to negotiate through regions of strong secondary structure, such as hairpins, often results in the formation of truncated PCR products. In addition, mis-priming amplification products are prevalent when primers are high in GC content. Despite the inherent challenges, detection of GC-rich sequences is becoming increasingly important for the molecular diagnosis of inherited diseases. Read More…
I was researching your CleanAmp primers and would like to know if they could be activated later in the PCR cycles?
I was researching your CleanAmp primers and would like to know if they could be activated later in the PCR cycles. For example, I have 2 pairs of primers for PCR and would like the first pair to work for cycle 1 through 10 followed by a long activation period to have both pairs working. What’s the probability that the 2nd pair would be activated during the first 10 denaturing cycles. – Chun-Nan Chen
Thank you for your question about CleanAmp Primers. For your interest in having one primer pair work earlier in PCR, with the second primer pair being activated after the first 10 thermal cycles, the best way to achieve this is to prepare the first primer pair without modifications and to prepare the second primer pair with CleanAmp Precision modifications… Read More
Have a question about CleanAmp™ Primers? Post a comment below.
I’m trying to conjugate some 2′-F-RNA aptamers onto nanoparticles. Can you recommend some functional linker that can be ligated to either end of the RNA and be used for conjugation to particles? – Johnson
Thank you for your question about how to best conjugate 2′-F-RNA aptamers onto nanoparticles. Depending on the type of nanoparticle you use (e.g., gold, quantum dot, magnetic or carbohydrate nanoparticles), there are different approaches for how to perform the conjugation… Read More
Have a question about aptamers? Post a comment below.
What is the Site-Directed Mutagenesis?
Site-directed or oligonucleotide-directed mutagenesis is a key method in the study of protein structure/function and gene expression control. If the nucleotide sequence of the gene is known, an oligonucleotide can be manufactured to introduce a single base change in the codon corresponding to the specific amino acid to be altered. The oligonucleotide is then used to generate a set of clones that can be identified and propagated. Learn More.
What is the use of a Trimer Oligo Solution?
While randomers are often used in protein mutagenesis procedures, they have several short comings. First, it is very difficult to achieve a truly random mixture. The second and more problematic issue is the inevitable introduction of stop codons. Fortunately, the ability to direct combinatorial mutagenesis using randomized oligonucleotides has been advanced by the use of trimer oligonucleotides. Read More.
Which RNA polymerase to use for incorporation of 2′ Fluoro and 2′ O-Methyl NTPs?
Wondering which RNA polymerase to use for incorporation of 2′ Fluoro and 2′ O-Methyl NTPs? We recommend T7 R&DNA™ and SP6 R&DNA™ polymerases from Epicentre. We’d like to hear from you – click the link below to leave your comment.
Have a Question?
Collectively the TriLink Team has many years of experience in nucleic acid chemistry to offer. Post your question in the comment field below and we will get an answer to you as soon as we can.
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Reprints of TriLink’s publications are available. To request a copy, please email info@trilinkbiotech.com.
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