Team:University of Lethbridge/Notebook/GeneralLabAugust

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Contents

August 13, 2008

Nathan Puhl and Sebastian

Verified and quantified the LacI and DT restriction digest products.

Set up a Ligase experiment for LacI and DT 

- 1.3 uL vector(DT)
- 8 uL insert(LacI)
- 2 uL 10x Ligase buffer
- 1.5 uL Ligase
- 7.2 uL ddH2O
- 20 uL total volume


August 14, 15

Nathan Puhl, Munima, Selina

Poured 61 LB + Amp plates. Stored in iGEM 4 C fridge.


August 16, 2008

Roxanne

Used Qiagen Plasmid MiniPrep Kit to Plasmid Prep Last Year's Biobrick Parts that were Incubated overnight.

Ran Plasmids in a 1% Agarose Gel at 100 V for 30 minutes.


August 18

Christa, Nathan Puhl, Munima

Ran a colony PCR of riboswitch extracted from gel done by Nathan and Roxanne on a previous day. Roxanne will remove the PCR tube from the thermocycler in the morning.


August 19, 2008

Christa, Munima, Sebastian

Did a plasmid prep on pSB1A7 using QIAprep Spin Miniprep Kit. Stocked 4- 50uL of pSB1A7 and stored in iGEM -20 C.

Sebastian, Nathan Puhl

Ran products from plasmid prep (pSB1A7 x 4 samples) on 1% agarose gel for 27 minutes. 

-Lane 1 - 1 kb GeneRuler ladder (2 uL)
-Lane 2 -6 pSB1A7 (3 uL) + 6x loading dye (2 uL); Mixed up what sample was in Lane 4, so Lanes 5 and 6 were run.

Conclusion: The bands appeared to be at the correct size for pSB1A7.


August 22, 2008

Christa, Munima, Nathan Puhl, Roxanne

Objective: Run a gel to confirm that appropriate inserts were amplified from the PCRs and do a gel extraction of the inserts to prepare them for the biobrick format. 

-Could not obtain a picture of the gel (1% agarose) of the digested pSB1A7 and the recently amplified CheZ gene because the camera would not turn on. 
  The CheZ gene appeared at the correct size (~700 bp).
-Did a gel extraction of pSB1A7 and CheZ from that gel with the Qiagen MiniElute Gel Extraction kit.
-Ran another 1% agarose gel to confirm that the gel extraction was successful. Bands appeared at appropriate sizes. The concentrations of pSB1A7 
  and CheZ were estimated to be 25 ng/uL and 80 ng/mL, respectively.

Next step: Digest both pSB1A7 and CheZ with phosphatase. A restriction digest will be performed at a later date on CheZ to prepare it for insertion.

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