User:Cassandra M Barrett/Notebook/Open Chromatin/2015/10/07

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LCR Plasmid Digest

Purpose: to check for any properly constructed plasmids via digesting sample and running it on a gel to check proper fragment size

Methods:

Using PstI FD, set up 11 digests: LCR colonies 1-8, purified MV10 from yesterday, sequence verified MV10 used for transformation, water

Create the following mastermix (X12)

  • 180uL H2O
  • 24uL 10X FD Buffer
  • 12uL FD PstI

Aliquot 18uL of mastermix and add 2uL of appropriate DNA.

Incubate at 37C for 15min.

Run samples on 1% agrose gel for 45min at 110V

Results:

  • Expected results for MV10: linear 5197bp
  • Expected results for MV10_Gal4DB_mCh_ATF2: 1814bp, 5459bp

All lanes appeared to be around 5kb like the sequence verified MV10 control. We can tell that the LCR didn't work and that the MV10 isolated from the + control is most likely the right thing.