Achieve high data quality from PCR products
ExoSAP-IT reagent may be used as an effective clean-up method prior to fluorescent or radioactive DNA sequencing, SNP analysis, or any other application requiring a PCR product free of excess nucleotides and primers.
The data below shows the results of downstream processing with PCR product cleaned up by ExoSAP-IT reagent, an alternative enzymatic reagent, or a spin column method. The ExoSAP-IT reagent data shows superior results with no miscalls or frame shift mutations.
Fig. 1. Fluorescent sequencing results of a 1,007 bp PCR product. A 1,007 bp fragment was amplified and
treated with ExoSAP-IT reagent (above) or an alternate enzymatic reagent (below) and sequenced. Pherograms
revealed no miscalls with ExoSAP-IT reagent but four miscalls at position 25, 26, 30, and 39 with the alternate
enzymatic reagent. 5 µl of PCR product was treated with 2 µl ExoSAP-IT reagent or the alternate
enzymatic method per its protocol. Note: Two of the four miscalls are frame-shifts.
Fig. 2. Fluorescent sequencing results of a 100 bp pUC18 PCR fragment sequenced with a -20 Fwd primer using fluorescent sequencing reagents. PCR clean-up performed with: (a) ExoSAP-IT reagent; (b) a column designed for PCR clean-up. Base miscalls in (b) are due to inherently low yields of short PCR products when using columns.
Recover 100% of your PCR product with USB ExoSAP-IT PCR Product Clean-Up:
the gold standard for enzymatic PCR clean-up.
- 100% sample recovery with one-tube, one-step PCR clean-up - no loss of PCR products, regardless of the fragment size
- Expertise from the original and only source of ExoSAP-IT reagent - the USB team has been producing, testing, and selling ExoSAP-IT reagent since 2000
- The gold standard in enzymatic PCR clean-up - ExoSAP-IT reagent has been referenced in thousands of publications since 2000; the market leader in enzymatic PCR clean-up with an established reputation for consistency and quality
- Quality results with SAP - only ExoSAP-IT reagent uses SAP, offering higher specific activity for better sequencing data when compared to alternative alkaline phosphatases
- Optimized formulation - the buffer formulation in ExoSAP-IT reagent has been optimized to give superior performance of the product
- Eliminate spin columns - decrease time and expense while increasing yield
- Degrades contaminating primers and dNTPs - no interference in downstream applications
- Green option - generate less waste with our single-tube solution to PCR purification vs. columns
- Sole source - Affymetrix is the only supplier of USB ExoSAP-IT reagent, ensuring your lab of product consistency and integrity
- Why lose time validating an alternate source - only ExoSAP-IT reagent has been around long enough to have validated stability measurements
ExoSAP-IT reagent is designed for simple, quick PCR clean-up for downstream applications, such as DNA sequencing or Single Nucleotide Polymorphism (SNP) analysis. When PCR amplification is complete, any unconsumed dNTPs and primers remaining in the PCR product mixture will interfere with these methods. ExoSAP-IT reagent removes these contaminants.
How it works
ExoSAP-IT reagent is added directly to the PCR product and incubated at 37°C for 15 minutes. After PCR treatment,
ExoSAP-IT reagent is inactivated simply by heating to 80°C for 15 minutes.
ExoSAP-IT single-step PCR clean-up utilizes two hydrolytic enzymes, Exonuclease I and Shrimp Alkaline Phosphatase, together in a specially formulated buffer, to remove unwanted dNTPs and primers from PCR products. Exo I removes residual single-stranded primers and any extraneous single-stranded DNA produced in the PCR. SAP removes the remaining dNTPs from the PCR mixture.
This method is designed to require a minimum of 'hands-on' time. Enzymatic removal of excess primers and unincorporated nucleotides occurs in one easy step by using ExoSAP-IT reagent in a single tube or microtiter well.
No sample loss
Use of the ExoSAP-IT method eliminates all gel or column purifications, sedimentations, filtrations, beads and/or magnetic separations. There is 100% recovery of both short and long PCR products with ExoSAP-IT reagent.
Use the ExoSAP-IT method for PCR Clean-Up and see the difference 100% can make.