Team:UCLA/Notebook/Spider Silk Genetics/22 July 2015

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7/22/2015

Transformation Results

• Colonies are present on all plates from yesterday.

Miniprep

• Used Zymo Classic kit to prep M2-9(T7) and M2-SeqAB

	Concentration (ng/uL)	A 260/280
M2-9(T7)-1	590.49	1.86
M2-9(T7)-2	614.07	1.84
M2-9(T7)-3	392.19	1.84
M2-Seq AB	1117.69	1.88

• Sent M2-9(T7) samples for sequencing using VF/R primers as needed.

BsaI Digestion for M2-Seq AB

• Digest 5 ug of M2-SeqAB plasmid in 50 uL reaction with 4 uL of BsaI.

	Volume (uL)
BsaI	4
10x Cutsmart	5
DNA	4.47
ddH2O	36.53

• Ran digest on 1.5% TAE gel. Used 2 uL of 50 bp ladder.

Fig. 1BsaI Digestion of MaSp2 Seq AB. The Expected product size is 102 bp.

• The digestion worked, but it is not as complete as previous digestions have been. Compare to previously.

Midi prep for M2-AB

• Used Qiagen Kit, eluted in 200 uL EB.
• Approximately 100 ng/uL for each)
• May need to only digest 4 ug of each in the future.

Bacterial Culture

• Picked three colonies from each plate from yesterday.
  • Grow in 5 uL liquid culture.
• Streaked one plate with all three of the M2-9(T7) samples we sent for sequencing.