Member since: 2019
Organization: MicrobiType LLC
The best way to prepare PCR products for Sanger sequencing!
Application Area: Sample preparation for sanger sequencing
"ExoSAP-IT is an enzyme mixture used to "clean up" PCR products (i.e., digest PCR primers) so they can be directly sequenced by the Sanger dideoxynucleotide method following addition of sequencing primer. It provides a simpler alternative to silica column clean up, and requires less hands-on time (although a half hour 37C-80C incubation in a thermal cycler is required). Note that the volumes of ExoSAP-IT and PCR product specified by the manufacturer can be scaled down 5-fold or so, making it very cost-effective (<$1 per sample). Most importantly, the sequence chromatograms are consistently excellent (depending on your PCR product, of course)."
ExoSAP-IT™ PCR Product Cleanup Reagent is used for enzymatic cleanup of amplified PCR product. It hydrolyzes excess primers and nucleotides in a single step. ExoSAP-IT-purified samples are ready for use in downstream applications such as DNA sequencing or single nucleotide polymorphism (SNP) analysis.
The ExoSAP-IT one-step solution provides a simple way to enzymatically purify a PCR reaction. The reaction setup is complete with one pipetting step, which is followed by two incubations. The first incubation digests excess primer and dephosphorylates nucleotides. The second, high-temperature incubation inactivates the enzymes.
Enzymatic cleanup is a very efficient method for PCR purification. Adding ExoSAP-IT reagent directly to the PCR product eliminates transfer steps to tubes, wells, or columns and conserves PCR amplicons, helping reduce the chance of cross contamination; no further processing needed.
After enzymatic cleanup with ExoSAP-IT reagent, primers and dNTPs will no longer interfere with DNA sequencing or SNP analysis.