Lewis' lab notebook

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Contents

6/14

  • 5 minipreps (R0010, E0241, E7104) Protocol used
  • R0010 and E0241 digests; digests were done according to Knight:Restriction_Digest; recipe halved.
  • Phosphatase was added to R0010 samples; protocol here.
  • Gel run to separate out DNA fragments resulting from R0010 and E0241 digests. 130V, 45 min.
  • Bands were almost invisible (wells used were those to the right of the second ladder). This may have been due to a second wash step during the mini-preps (although this is unlikely).

Image:Zs_jl_lh_pt_6-14_digest.jpg

6/15

6/19

  • Selected colonies containing R0010+E0241 fragments, which appeared to be constitutively on. For more details, ask Nick.
  • Cultured two colonies in 5mL LB + 50uL Amp (5mg/uL orig) overnight @37 on shaker.

6/20

  • Made a glycerol stock of two colonies.
  • Minipreps carried out. This protocol was used except final elution in 30 uL water to increase DNA concentrations.
  • Digests were done according to Knight:Restriction_Digest; recipe halved. REs = XbaI, PstI.
  • 1.2% E-gel run using 10 uL digest + 10 uL water per well. See lanes 5 and 6 in image below.

Image:Digest 6-20-06.jpg

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