Team:Alberta/Notebook/protocols/ligation

Ligation

Reagents:

  • Vector
  • Insert
  • MilliQ buffer
  • 5X ligase buffer
  • T4 DNA ligase
    • [http://tools.invitrogen.com/content/sfs/manuals/t4dnaligase_5U_man.pdf]


Procedure:

  • Volumes of vector and insert may vary, but a typical 20 ul reaction might be:
MilliQ water 11 ul
5x ligase buffer 4 ul
Vector (200 ng/ul) 1 ul
Insert (200 ng/ul) 3 ul
Ligase 1 ul
  • Incubate at R/T for 1 hour or overnight at 12-16oC.


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