Matt:JustinLabNotes/2015-10-14
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CircLigase II Test Reagents[edit]
- CircLigase II kit is on back order
- I have plenty of enzyme but low on 10X buffer, 5M Betaine, and 50mM MnCl2
Reagents[edit]
Justin made from dry reagent[edit]
- 5M Betaine
- 50mM MnCl2
Neither worked as well as kit-provided reagents
Ordered from 3rd party companies[edit]
- 5M Betaine from SigmaAldrich
- 50mM MnCl2 from Trevigen
Results[edit]
File:CircLigase ii Reagents test - 10-14-15.jpg
- Even positive control didn't circularize
- I believe the second band is 5' adenylation from the enzyme and not circularization since in the epicentre protocol the circularized band runs slower than 70bp ladder
- Also the circularized band showed up in Justin's gel last week
- Try ExoI digestion to ensure