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| - | Protocol 14: Fluorescent Sequencing Reaction
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| - | Procedure:
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| - | *In a 0.2ml PCR tube, add:
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| - | {|
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| - | |Template || 5.0ul (200 ng/ul)
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| - | |-
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| - | |VF primer || 1.0ul
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| - | |-
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| - | |Dilute buffer || 2.5ul
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| - | |-
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| - | |BD sequence mix || 1.5ul
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| - | |}
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| - | * Mix well.
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| - | * Select program 'seq-dye' on PTC 200 thermal cycler.
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| - | * Run program. It will take about 2 hours.
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| - | * Remove tube from PCR machine and transfer 10ul rxn mix to 1.5ml Eppendorf tube. Add:
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| - | |Blue NaOAc/EDTA || 1.5ul
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| - | |-
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| - | |95% ethanol || 40ul
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| - | |}
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| - | * Let sit on ice 15 minutes.
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| - | * Centrifuge 10 minutes at max speed.
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| - | ** Should see a small blue dot at bottom of tube.
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| - | * Discard supernatant.
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| - | * Wash pellet with 500ul of 70% ethanol.
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| - | * Discard supernatant and spin briefly to bring down the residual liquid. Draw liquid off with a P10 pipette tip.
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| - | ** Do not disturb the pellet.
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| - | * Air dry for 10 minutes.
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| - | * Place in -20<sup>o</sup>C freezer to be delivered to MBSU 4th floor microbiology M534. Reactions delivered before 2pm will normally be returned the next day.
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| - | [[Team:Alberta/Notebook/protocols| Back]]
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| - | {{Team:Alberta/endMainContent}}
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