Daniel:Notebook/ComboLock/2017-3-3
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Padlock0601 Test (Started Yesterday)
Protocol-Part 5
- TBE Gel
- Mix 240 uL TBE, 60 uL 6x loading dye
- Aliquot 10 uL per sample/ladder lane onto parafilm
- Add 2 uL of sample or ladder to correct drop
- Load 10 uL in to well
- Run gel for 23 minutes at 230V
- Open gel and stain with 3 uL SYBR Gold for 3 minutes
- Rinse gel and image in gel doc
- qPCR
- Make 9.2X qPCR master mix according to following recipe
- 193.2 uL nfH2O
- 230 uL SYBR Fast
- 9.2 uL 10 uM AmpF
- 9.2 uL 10 uM AmpR-Ind20
- Add 48 uL master mix to each well
- Add 2 uL sample according to plate layout File:PlateLayout-20170303-Padlock0601-Bglii.png
- qPCR Cycles
- 95C 3 min
- 95C 3 sec
- 55C 30 sec
- 72C 20 sec
- plate read
- goto b x30
- 72C 2 min
- 16C hold
- TBE Gel
- Mix 96 uL TBE, 24 uL 6x loading dye
- Aliquot 10 uL per sample/ladder lane onto parafilm
- Add 2 uL of sample or ladder to correct drop
- Load 10 uL in to well
- Run gel for 23 minutes at 230V
- Open gel and stain with 3 uL SYBR Gold for 3 minutes
- Rinse gel and image in gel doc
Results
- 20170302-qPCR-Padlock0601Test.png
qPCR curves for run on 3-2-2017
- 2017-03-03-Padlock0601-PreRCA-Unpurifed.png
Gel image-PreRCA Unpurified
- 2017-03-03-Padlock0601-PreRCA-Purifed.png
Gel image-PreRCA Purifed
- 2017-03-03-Padlock0601-PostRCA-Unpurifed.png
Gel image-PostRCA Unpurified
- 2017-03-03-Padlock0601-PostRCA-Purifed.png
Gel image-PostRCA Purified
- 2017-03-03-Padlock0601-Bglii.png
Post BglII digestion