Matt:LabNotes/2014-2-6: Difference between revisions

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>Mzcai
>Mzcai
 
Line 20: Line 20:
  Program stored under Kun/CPG
  Program stored under Kun/CPG
  -> 95c 30sec -> cool down to 55C at 0.02C/sec -> 55C 16.5h (started at 6pm)
  -> 95c 30sec -> cool down to 55C at 0.02C/sec -> 55C 16.5h (started at 6pm)
  -> add 3ul ampligase mix (0.5U/ul AmpLigase)
  -> add 3ul ampligase mix (0.5U/ul AmpLigase in 1X Buffer)
  -> 55C 5h-> 94C 2min -> add 2ul Exo I/III mix-> 37C 2h -> 94C 2min -> 4C hold.
  -> 55C 5h-> 94C 2min -> add 2ul Exo I/III mix-> 37C 2h -> 94C 2min -> 4C hold.


*Stored in -20C until purification
*Stored in -20C until purification

Latest revision as of 18:28, 21 February 2014

ppMALAT1 Capture in vitro[edit]

  • Will do 8 reactions of 35ul, so almost 19x more
' 100uM MALAT1 Template H2O 200uM ppMALAT1 10X Ampligase Total
1 Reaction 6 18 3 3 30
8 Reactions 48 144 24 24 240
  • Chose to use more template and less padlock probe than Alan because we want all padlock probes to circularize
    • So more like 12x more but hopefully with higher yield
Program stored under Kun/CPG
-> 95c 30sec -> cool down to 55C at 0.02C/sec -> 55C 16.5h (started at 6pm)
-> add 3ul ampligase mix (0.5U/ul AmpLigase in 1X Buffer)
-> 55C 5h-> 94C 2min -> add 2ul Exo I/III mix-> 37C 2h -> 94C 2min -> 4C hold.
  • Stored in -20C until purification