User:Behzad Damadzadeh/Notebook/PcTF Subcloning in E-coli/2013/05/06: Difference between revisions
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*Ligation of construct BD001(KAH182-KAH201) and mammalian vector V0200 and cloning in Bl21-DE3. | *Ligation of construct BD001(KAH182-KAH201) and mammalian vector V0200 and cloning in Bl21-DE3, The new construct is named: BD002. | ||
*KAH182 = BBa_S04739 and KAH201 = BBa_S04745 and V0200 = BBa_J176121. | *KAH182 = BBa_S04739 and KAH201 = BBa_S04745 and V0200 = BBa_J176121. | ||
*Assembly strategy: The backbone V0200(N/S)/4442 , the insert is BD001 (KAH182-KAH201) (N/S)/1901. | *Assembly strategy: The backbone V0200(N/S)/4442 , the insert is BD001 (KAH182-KAH201) (N/S)/1901. | ||
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| Ligation || <font color="blue"><u> | | Ligation || <font color="blue"><u> | ||
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| 1. E. coli BL21 | | 1. E. coli BL21 BD001 (KAH182-KAH201) (N/S)/size, 21.00 ng + V0200(N/S)/size, 43.00ng || <font color="blue">KAH182/KAH201 3:1 Six Colonies</font> | ||
|- | |- | ||
| 2. E. coli BL21 KAH182-KAH201(N/S)/size, 21.00 ng + V0200(N/S)/size, 43.00ng || <font color="blue">KAH182/KAH201 4:1 One Colony</font> | | 2. E. coli BL21 BD001 (KAH182-KAH201)(N/S)/size, 21.00 ng + V0200(N/S)/size, 43.00ng || <font color="blue">KAH182/KAH201 4:1 One Colony</font> | ||
|- | |- | ||
| 3. E. coli BL21 KAH182-KAH201(N/S)/size, 21.00 ng + V0200(N/S)/size, 43.00ng || <font color="blue">KAH182/KAH201 5:1 One Colony</font> | | 3. E. coli BL21 BD001 (KAH182-KAH201) (N/S)/size, 21.00 ng + V0200(N/S)/size, 43.00ng || <font color="blue">KAH182/KAH201 5:1 One Colony</font> | ||
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| 4. E. coli BL21 KAH182-KAH201(N/S)/size, 21.00 ng + V0200(N/S)/size, 43.00ng || <font color="blue">KAH182/KAH201 10:1 One Colony</font> | | 4. E. coli BL21 BD001 (KAH182-KAH201)(N/S)/size, 21.00 ng + V0200(N/S)/size, 43.00ng || <font color="blue">KAH182/KAH201 10:1 One Colony</font> | ||
|- | |- | ||
| 5. | | 5. V0200 (N/S)/size, 43.00ng (Negative Control Plate)|| <font color="blue">No Colony</font> | ||
|} | |} | ||
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'''Confirm The Assembly''' | '''Confirm The Assembly''' | ||
[[Image: | [[Image:Ligation Confirmation.jpg|thumb|350px| Two seperate bands in each column show the vector backbone and the insert, 4442 bps and the BD001 (KAH201+KAH182) (1901) constructed gene.Backbone, 1901 bps, Cut with SpeI and NotI. First column: Ladder, columns 1-5 are 3:1 ratio colonies, column6 is 4:1 ratio, column 7 is 5:1 ratio and column 8 is 10:1]] | ||
{| {{table}} cellspacing="3" width=400px | {| {{table}} cellspacing="3" width=400px | ||
| Plasmid DNA || 2.0 μl | | Plasmid DNA || 2.0 μl |
Revision as of 13:42, 6 May 2013
PcTF Subcloning in E. coli | <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page <html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
06/05/2013Building a Reporter Gene to Determine Chromatin Protein Function (Ligation into the mammalian vector)
Confirm The Assembly
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