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USTC/Notebook/DNA ligation
From 2008.igem.org
DNA Ligation
DNA ligases are used with restriction enzymes to insert genes into plasmids.
Materials
- DNA plasmids double digested previously(eg.XbaI and PstI); . Measure concentration in Nanodrop beforehand.
- Biobricks double digested previously(eg.SpeI and PstI )
- equal amount of T4 ligase.
Procedure
The following volumes apply to a 20µl DNA ligation.
add the following components to PCR tube:
- 9µl DNA plasmids
- 1µl biobricks
- 10ul Solution I (T4 ligase and buffer)
- MIX the reaction by stiring
- Incubate the reaction at 16*C for 2hrs or overnight to ensure complete ligation.
