Because the purity of the primer cannot be 100%, it is possible to select a clone that has been amplified by the impurity primer during the selection of the clone. At this time, please re-select a clone for sequencing, and you will get the correct result. If the sequencing results of 2 ~ 3 clones have not improved, we will re-synthesize the primer for free and deliver it to you as soon as possible. If you file a claim, according to international industry practice, the claim scope is limited to the price of the product.

EB stains DNA by intercalating between the base pairs of a double helix. Synthesized DNA molecules are single-stranded and can only be stained by EB if they form local hairpin loops or partial double-stranded structures by folding back on themselves. Since the sequences of different primers are different, their ability to form double helices is also different. Therefore, the brightness of the EB-stained bands cannot be used to quantify the synthesized DNA.

PCR reactions can fail for a variety of reasons. Here are some of the most common causes:Primer-template annealing: The primers must be complementary to the template sequence in order to anneal properly. If the primers do not anneal, the PCR reaction will not be able to start.Primer structure: The primers should not have any secondary structures that could interfere with annealing.Reagent quality: The PCR reagents must be of high quality in order to function properly.PCR instrument: The PCR instrument must be working properly in order to generate the desired results.PCR reaction conditions: Th…

For primer electrophoresis, it is essential to employ denaturing PAGE electrophoresis. As the primers are single-stranded DNA and prone to forming intricate three-dimensional structures, performing Agarose gel electrophoresis can lead to the appearance of multiple bands, making it impractical for quantitative analysis.

Because nucleic acids exhibit strong absorption near 260nm, and proteins show strong absorption near 280nm, the A260/A280 ratio is commonly used to assess nucleic acid purity when extracted from biological samples, with the ratio typically falling between 1.8 and 2.0. This judgment is based on the assumption that the proportions of A, G, C, and T in the sequence are roughly equal. However, for synthesized DNA/RNA, especially with short sequences (usually between 20 and 30 bases), the proportions of A, G, C, and T can vary significantly. Due to the different molar extinction coef…

No, both the 5' and 3' ends have -OH groups. If a phosphate group is needed, modification synthesis is required to introduce phosphorylation.

A convenient method in the laboratory is using the PAGE (Polyacrylamide Gel Electrophoresis) technique. Electrophoresis is performed on a polyacrylamide gel with 7M urea at a specific concentration. For primers with <12 bases, use a 20% gel; for primers with 12-60 bases, use a 16% gel; for primers with >60 bases, use a 12% gel. Take around 0.2 OD of the primer and dissolve it in urea-saturated solution or add urea powder to the primer solution until saturated. Before loading, heat denature the sample (95°C, 2 mins). The addition of urea serves the purpose of denaturation and increases s…

Typically, when synthesizing long-chain primers, the amount of reagents required is significantly higher than for short-chain primers, especially for primers exceeding 90 base pairs. Due to the increased cost of reagents, this results in an overall higher price for the synthesis of long-chain primers.

Sequences up to 150 bases in length can be synthesized.

The rough calculation for the nmol of synthesized oligo is done using the formula:\text{nmol} = \frac{\text{OD value} \times 33 \mu g}{\text{Number of bases} \times 330} \times 1000 = \frac{\text{OD value}}{\text{Number of bases}} \times 100nmol=Number of bases×330OD value×33μg×1000=Number of basesOD value×100For an accurate concentration of synthesized DNA, you can use the formula:\text{nmol} = \frac{\text{OD value} \times 1000}{(15.3 \times A) + (7.4 \times C) + (11.8 \times G) + (9.3 \times T)}nmol=(15.3×A)+(7.4×C)+(11.8×G)+(9.3×T)OD value×1000Where A, C, G, and T represent the re…

Oligos can be quantified using a UV spectrophotometer by measuring the absorbance of the solution at a wavelength of 260nm. It is recommended to dilute the solution to an absorbance range of 0.2-0.8 during measurement (extremely high or low absorbance can lead to significant errors). For DNA powder, it is dissolved in water with thorough shaking, and a portion of the solution is diluted to 1ml for measuring absorbance in a 1ml standard cuvette. The measured OD value for the given volume can then be used to calculate the OD value of the original solution.

Unresolved primers are highly stable and can be stored for at least one year at -20°C. For dissolved primers, it is recommended to pre-dilute them to a storage solution of 100 μM, aliquot into tubes, and store at -20°C. This can preserve them for at least six months or more (repeated freeze-thaw cycles can reduce their shelf life). Before use, dilute the concentrated solution to the working concentration for experiments.

Our synthesis report provides the amount of water to add for diluting each OD of the primer to a concentration of 100 μM (or 100 pmol/μl). For dissolution, you can add an appropriate volume of RNase-free double-distilled water (pH > 6.0) or TE buffer (pH 7.5-8.0) based on your experimental needs. Before opening the vial, it is advisable to centrifuge it at a speed of 3000-4000 rpm for 1 minute to prevent primer loss when the cap is opened.

Purification methods for Aegi primers:1. **C18 Column Desalting:** Also known as a simple reverse-phase column, it has specific adsorption for DNA, which can be eluted with an organic solvent but not with water. This effectively removes salt.2. **R-PAGE (Reverse-Phase PAGE):** Based on specific or reverse chromatography, it removes failed sequences from the synthesized product, suitable for primers below 35 bases.3. **PAGE Purification:** Using denaturing polyacrylamide gel electrophoresis, it separates primer DNA and then recovers the target DNA from the gel.4. **HPLC Purification:** Using th…

Primers are synthesized using the solid-phase phosphoramidite method. Various DNA synthesizers exist, but regardless of the machine used, the synthesis principles remain the same. The main differences lie in the synthesis yield, reagent consumption, and the time taken for a single cycle.1. **Deprotection:** Addition of deblocking reagent to remove the protecting group (DMT) from the 5'-OH of the base, obtaining a free 5'-OH.2. **Coupling:** Simultaneous addition of an activator and a new nucleotide. The 5'-OH of the new base remains protected by DMT, and a coupling reaction occurs …