Rough Timing Guide of Lab Training Session

From 2006.igem.org

Rough Timing Guide of Lab Training Session

This is an outline of preparatory activities that each group is expected to perform while in training. This outline is general, and the timeline will vary with each group’s performance.

Before Training Session Begins

The Operational Committee will prepare the following:

• Making of competent cells

o Take colony from plate, put in prepared broth, grow over night (O/N)

o In morning, take growth, thaw at 0oC, and purify cells ≈1hr

Training Session 1 (≈1.5-2.0 hrs)

The Training Group will complete the following tasks:

• Professor Davies safety tour ≈20 min

• Perform DNA extraction ≈70 min involving:

o Take tray of DNA

o Identify two parts needed

o Re-suspend DNA in pipette with buffer

o Mix with competent cells

o Incubate at 37oC ≈50 min

o Spread on petri dish with antibiotic resistance O/N

Before Next Training Session Begins

The Operational Committee will prepare the following:

• Take colonies that grew on the petri dish from transformation done by group during last training session, place in broth, and grow O/N

Training Session 2 (≈1.5-2.0 hrs)

The Training Group will complete the following tasks:

• With the cells that grew O/N (prepared by organizational committee), take about 3ml and place in microcentrifuge tubes

• Add glycerol and store in fridge for when we need the cells again

• With remaining O/N culture, perform plasmid extraction using specified kit (miniprep) ≈1 hr

Training Session 3 (≈4.0-5.0 hrs)

The Training Group will complete the following tasks:

• Prepare two separate tubes with restriction enzyme. One will have needed vector, while the other the purified plasmid. Let enzyme digest ≈1 hr

• Prepare gel for electropheresis

• Load both samples (digests) on gel, and run for ≈40-60 min

• Physically cut out (with razor blade) desired biobrick part and place in new test tube

• Use gel extraction kit to purify segments ≈1 hr

• You know have sticky ends, therefore place desired parts in same test tube with ligase to do ligation ≈15-40 min (varies on protocol)

• With product of ligation, add to competent cells, and perform transformation as before

--Jovan 22:46, 14 June 2006 (EDT)

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