Magni:Protocols/Digestion

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(New page: {{Magni}} <div style="padding: 10px; width: 720px; border: 5px solid #000000;"> = Digestion = To perform a double digest with the Roche enzymes, add to a 0.2-ml PCR tube: **1000 ng of DNA...)
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**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
**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
**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 or up to 20.5 ul of DNA (if performing a screening)
+
**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.
*Milli-Q water up to 20.5 ul.
*2.5 ul of buffer H
*2.5 ul of buffer H

Revision as of 13:38, 22 September 2012

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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 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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