Kun:LabNotes/ExonomeSeq/2007-12-20: Difference between revisions

From ZhangLabWiki
Jump to navigation Jump to search
(New page: ==Exp.1 Cloning of the size selected fragments from 12/19/07== ===Blunting of the DNA fragment=== 5X Phusion buffer: 1ul 10mM dNTP: 0.25ul DNA: 3...)
 
 
(2 intermediate revisions by the same user not shown)
Line 1: Line 1:
==Exp.1 Cloning of the size selected fragments from 12/19/07==
==Exp.1 Cloning of the size selected fragments from 12/19/07==
    PAGE size selected fragments:
    [[Image:2007-12-20-E55kv1-PAGE-selected-fragments.png|500px]]
    DNA concentration:
    GC1EP:    5.36ng/ul
    H2C:      1.64ng/ul
    150bp:    3.73ng/ul
    180/280bp  2.64ng/ul
    350bp:    5.17ng/ul
    470bp:    too low to measure
===Blunting of the DNA fragment===
===Blunting of the DNA fragment===
       5X Phusion buffer:  1ul
       5X Phusion buffer:  1ul
Line 13: Line 22:
     T4 DNA ligase (4U/ul) 0.5ul
     T4 DNA ligase (4U/ul) 0.5ul
     16C 1h -> 65C 10min -> 4C hold.
     16C 1h -> 65C 10min -> 4C hold.
===Colony PCR===
  [[Image:2007-12-23-Colony-PCR.png|300px]]

Latest revision as of 02:39, 24 December 2007

Exp.1 Cloning of the size selected fragments from 12/19/07[edit]

   PAGE size selected fragments:
   File:2007-12-20-E55kv1-PAGE-selected-fragments.png
   DNA concentration:
   GC1EP:     5.36ng/ul
   H2C:       1.64ng/ul
   150bp:     3.73ng/ul
   180/280bp  2.64ng/ul
   350bp:     5.17ng/ul
   470bp:    too low to measure

Blunting of the DNA fragment[edit]

      5X Phusion buffer:   1ul
      10mM dNTP:        0.25ul
      DNA:              3.65ul
      Phusion:           0.1ul
      72C 10min

Blunt-end ligation[edit]

    pCR-Blunt             0.5ul
    Blunt PCR product       2ul
    10X Ligation buffer   0.5ul
    Sterile water         1.5ul
    T4 DNA ligase (4U/ul) 0.5ul
    16C 1h -> 65C 10min -> 4C hold.

Colony PCR[edit]

 File:2007-12-23-Colony-PCR.png