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PcTF Subcloning in E. coli

01/12/2013

Ligation of KAH111, KAH112 and C-His vector in DH5α-T and BL21

The 4 μL of C-His Vector from stock is diluted with 16 μL of nuclease free water, then cut with E/P and purified from the gel.
Ligations

Ligation	</font>
1. E. coli BL-21 KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng	KAH111/C-His 3:1 No Colonies
2. E. coli DH5α-T KAH111(E/P)/size, 16 ng + C-His(E/P)/3600, 26ng	KAH111/C-His 3:1 No Colonies
3. E. coli BL-21 KAH112(E/P)/size, 22.8 ng + C-His(E/P)/3600, 26ng	KAH112/C-His 3:1 No Colonies
4. E. coli DH5α-T KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng	KAH112/C-His 3:1 No Colonies
5. C-His(E/P)/ 26 ng (Control Plate)

Calculations are for the ng of insert we need to get a 3:1 ratio of insert molecules to 50 ng vector molecules

Revision as of 17:13, 12 January 2013 (view source) Behzad Damadzadeh (Talk \| contribs) (→01/12/2013) ← Previous diff		Current revision (17:52, 12 January 2013) (view source) Behzad Damadzadeh (Talk \| contribs) (→01/12/2013)
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	* The incubation time for the ligation process was 30min at room temperature.		* The incubation time for the ligation process was 30min at room temperature.
		+	* Fast transformation, 30 min on ice.