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Services – ExoSAP-IT™ Express

• ExoSAP-IT™ Express reagent

• ExoSAP-IT vs. EtOH-EDTA precipitation

• Online Submission Request

• Physical Submission of Samples

ExoSAP-IT™ Express reagent

Before sequencing PCR products, residual primers and unincorporated nucleotides must be removed from the samples. To accomplish this, this product uses two hydrolytic enzymes: a modified Exonuclease I; and, Shrimp Alkaline Phosphatase [SAP]. For full product details, visit the ThermoFisher website: ExoSAP-IT Express [A35004]).

In brief, ExoSAP-IT is active in commonly used PCR buffers, so no buffer exchange is required and the reaction occurs in a single tube with just one pipetting step. The reagent is added directly to the PCR product for a 4 minute cleanup at 37oC, followed by a 1 minute inactivation at 80oC; an inert tracking dye offers visual confirmation that the reagent has been added. Finally, none of the kit components will interfere with downstream applications.

ExoSAP-IT vs. EtOH-EDTA precipitation

There are pros-&-cons to using ExoSAP-IT Express vs. the Core's 'primer-depletion' EtOH-EDTA precipitation protocols.

  • Pro: This kit retains 100% of the original PCR product... whereas one can expect to lose some (usually <20-30%) of the PCR products in the EtOH-EDTA procedures. [However, one might still be using the same amount of PCR product because only ~50% of the treated sample is used in a sequencing reaction.]
  • Pro: Because ExoSAP-IT retains 100% of dsDNA products, it can be used as a troubleshooting tool when there are concerns that the EtOH-EDTA protocols are losing too much PCR product.
  • Con: Unlike the EtOH-EDTA primer-depletion protocols, ExoSAP-IT will not remove primer-dimers or other non-specific small PCR products. If present in the sample, such small non-targets will create noise in the early sequence data.
  • Con: Unless doing just a few samples, it is much more expensive to use this kit.
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    Online Submission Request

  • Log In to the website, and click on "Services Page":
    • Make the necessary 'selections'.
    • In the "Client Memo" field, specify ...
    • Click the 'check box', and submit your request.
  • If you make an error in your request, you may revise that request if you do so before we begin processing it.
  • You will receive an automated email when your samples are processed and ready for pick-up.
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    Physical Submission of Samples

  • Tubes:  Samples exceeding ~25 ul should be submitted in 0.5-1.5 ml tubes (standard or Eppendorf LoBind); otherwise, 0.2-ml tubes are acceptable.
  • Label Submission:  Upon successful submission of your online request, the 'Submission #" will appear in your 'Submission History'. Then, label the tube rack itself with:
    • Submission #;
    • Date;
    • PI's Last Name; and,
    • Submitter name or initials.
  • Delivery site:  Transfer tubes to a Genomics Core rack (taking your own rack back to your lab), and put samples in the Mini-fridge (by sink) in the Genomics Core (A628 Life Science Annex).

    [Note: If the Core is locked, put your samples in the plastic "After-Hours" GC drop-box located in the adjacent Cold Room (beside sink, Rm. A650).]

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