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Protocol/8
From 2010.igem.org
(Difference between revisions)
| Line 14: | Line 14: | ||
| - | + | Reagents: | |
* Vector | * Vector | ||
* Insert | * Insert | ||
| - | * | + | * MilliQ buffer |
* 5X ligase buffer | * 5X ligase buffer | ||
* T4 DNA ligase | * T4 DNA ligase | ||
** [http://tools.invitrogen.com/content/sfs/manuals/t4dnaligase_5U_man.pdf] | ** [http://tools.invitrogen.com/content/sfs/manuals/t4dnaligase_5U_man.pdf] | ||
| - | Volumes of vector and insert may vary but a typical 20 ul reaction might be | + | |
| + | 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 | | 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 | + | Incubate at R/T for 1 hours or overnight at 12-16<sup>o</sup>C |
| + | |||
[[Team:Alberta/Notebook/protocols| Back]] | [[Team:Alberta/Notebook/protocols| Back]] | ||
{{Team:Alberta/endMainContent}} | {{Team:Alberta/endMainContent}} | ||
Revision as of 01:19, 26 October 2010
Protocol 8: 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 hours or overnight at 12-16oC
