Matt Gethers/CRI, Thailand/Labwork/Digests/NdeI and ApaI Double Digest

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NdeI and SacI Double Digest

Reaction Mix

Reagent Volume
Template 10 μl
NdeI 0.5 μl
SacI 0.5 μl
Fast Digest Buffer 2 μl
H20 7.0 μl
Total 20 μl

Reaction Conditions

Incubate mixture at 37oC for 30 minutes and inactivate at 65oC for 15 minutes on the heat block.

Run Notes

7.29.08

Used to digest pCM351 to excise Lox/Gentamcin cassette. 2.5 μl pCM351, 0.5 μl both enzymes, 2 μl buffer, and 14.5 μl water. Incubated at 37 degrees for 1 hour, 15 minutes, inactivated at 65 degrees for 15 minutes. Ran out on gel and excised ~1 Kb band. Did a gel purification on the morning of 7.30.08 and eluted in 50 μl elution buffer.

8.12.08

Digested pCM351 to excise Lox/Gentamcin cassette. 2.5 μl pCM351, 0.5 μl both enzymes, 2 μl buffer, and 14.5 μl water. Incubated at 37 degrees for 30 minutes, inactivated at 65 degrees for 15 minutes. Ran out on gel and excised ~1 Kb band. Did a gel purification and eluted in 50 μl elution buffer.

References

NdeI

SacI

Double Digest