Magni:Protocols/Digestion

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(Digestion)
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*1 ul of the second enzyme
*1 ul of the second enzyme
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Mix well and incubate in the thermocycler (''Digestion'' program) for 2-4 hours if using the products for a ligation, or for 10-20 minutes if using the products for a screening.
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Mix very well and incubate in the thermocycler (''Digestion'' program, reaction volume of 25 ul) for 1-4 hours if using the products for a ligation, or for 10-20 minutes if using the products for a screening.

Revision as of 13:35, 2 February 2013

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Digestion

To perform a double digest with the Roche enzymes, add to a 0.2-ml PCR tube:

    • 1000 ng of DNA or up to 20.5 ul of DNA (if performing the excision of an insert for a ligation, e.g. E-P, X-P, E-S) or
    • 500-700 ng of DNA or up to 20.5 ul of DNA (if opening a plasmid E-X or S-P for a ligation) or
    • 50-1000 ng of DNA (200 ng is ok for most cases) or up to 20.5 ul of DNA (if performing a screening)
  • Milli-Q water up to 20.5 ul.
  • 2.5 ul of buffer H
  • 1 ul of the first enzyme
  • 1 ul of the second enzyme
Mix very well and incubate in the thermocycler (Digestion program, reaction volume of 25 ul) for 1-4 hours if using the products for a ligation, or for 10-20 minutes if using the products for a screening.
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