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| - | == Rough Timing Guide of Training Session ==
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| - | 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.
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| - | '''''Before Training Session Begins'''''
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| - | The Operational Committee will prepare the following:
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| - | • Making of competent cells
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| - | o Take colony from plate, put in prepared broth, grow over night (O/N)
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| - | o In morning, take growth, thaw at 0oC, and purify cells ≈1hr
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| - | '''''Training Session 1 (≈1.5-2.0 hrs)'''''
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| - | The Training Group will complete the following tasks:
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| - | • Professor Davies safety tour ≈20 min
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| - | • Perform DNA extraction ≈70 min involving:
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| - | o Take tray of DNA
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| - | o Identify two parts needed
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| - | o Re-suspend DNA in pipette with buffer
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| - | o Mix with competent cells
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| - | o Incubate at 37oC ≈50 min
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| - | o Spread on pitre dish with antibiotic resistance O/N
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| - | '''''Before Next Training Session Begins'''''
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| - | The Operational Committee will prepare the following:
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| - | •Take colonies that grew on the petri dish from transformation done by group during last training session, place in broth, and grow O/N
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| - | '''''Training Session 2 (≈1.5-2.0 hrs)'''''
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| - | The Training Group will complete the following tasks:
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| - | •With the cells that grew O/N (prepared by organizational committee), take about 3ml and place in microcentrifuge tubes
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| - | •Add glycerol and store in fridge for when we need the cells again
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| - | •With remaining O/N culture, perform plasmid extraction using specified kit (miniprep) ≈1 hr
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| - | '''''Training Session 3 (≈4.0-5.0 hrs)'''''
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| - | The Training Group will complete the following tasks:
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| - | •Prepare two separate tubes with restriction enzyme. One will have needed vector, while the other the purified plasmid. Let enzyme digest ≈1 hr
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| - | •Prepare gel for electropheresis
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| - | •Load both samples (digests) on gel, and run for ≈40-60 min
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| - | •Physically cut out (with razor blade) desired biobrick part and place in new test tube
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| - | •Use gel extraction kit to purify segments ≈1 hr
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| - | •You know have sticky ends, therefore place desired parts in same test tube with ligase to do ligation ≈15-40 min (varies on protocol)
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| - | •With product of ligation, add to competent cells, and perform transformation as before
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