IGEM:UBC/2009/Notebook/UBC iGEM 2010/2010/06/25

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Transformations
Protocol: See Transformation Protocol in "iGEM common protocols". It is written below for easy access. Supplies Needed: Competent cells Water bath/heat block at 42°C Ice SOC/LB medium LB agar plates (with appropriate selection)

Steps:
 * 1) Remove competent cells (100uL aliquots) from -80°C and thaw on ice.
 * 2) Add 1µL ligation mix to thawed cells and incubate for 30 minutes on ice.
 * 3) Heat shock in water bath/heat block for 60 seconds and put back on ice for two minutes.
 * 4) Add 400uL LB medium and incubate at 37C for 2 hours.
 * 5) Spread plate entire 500µL.
 * 6) Incubate overnight (<18hrs) at 37°C.


 * Changes:
 * Add 10uL ligation mix
 * Spread on Chloramphenicol plates
 * Used competent cells: DH5&alpha;

*Note: Could not add 10uL to 10T because not enough (see previous day)
 * Chose samples 5,6,11,12 because H171 showed distinct bands on the gel verification of the PCR products
 * Chose samples 2,4,8,10 at random - one from each strain: H49 and H80 (of different primers: one with His6-tag and without His6-tag)

Incubate @ 37°C starting 1340 Took out of incubator at 1541

Plated on Chloramphenicol plates - spread 500uL for all Began incubation @ 37°C at 1608 Vicki Ma 04:58, 28 June 2010 (EDT)
 * Note: Did not fully spread 500uL on 4T (did not have enough for some reason - lacking 0.5uL to 1uL)