Difference between revisions of "User:Behzad Damadzadeh/Notebook/PcTF Subcloning in E-coli/2012/11/16"

From OpenWetWare
Jump to: navigation, search
(11/16/12)
(11/16/12)
Line 32: Line 32:
 
|-valign="top"
 
|-valign="top"
 
| <u>Reagent</u> || <u>Volume</u> || &nbsp;
 
| <u>Reagent</u> || <u>Volume</u> || &nbsp;
| rowspan="9" | [[Image:KAH_060311_gel1.tif|350px|Cloning digests 6/03/11]]<br>30 μL/lane, 1% agarose
+
| rowspan="9" | <!--- [[Image:image.tif|350px|Cloning digest ]]<br>30 μL/lane, 1% agarose --->
 
|-
 
|-
 
| DNA (plasmid) || 25.0 μL
 
| DNA (plasmid) || 25.0 μL

Revision as of 11:55, 16 November 2012

Owwnotebook icon.pngPcTF 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>

11/16/12

  • ---Karmella 13:51, 16 November 2012 (EST): Here is my suggested procedure for trying the PcTF fusion cloning...

Assemblies: Shown as Part/cuts/purified fragment size

  1. KAH109/pT7CFE1-CHis: KAH109/(E/P)/1132 + pT7CFE1-CHis/(E/P)/3600
  2. KAH111/pT7CFE1-CHis: KAH184/(E/S)/1123 + "
  3. KAH112/pT7CFE1-CHis: KAH187/(E/S)/1123 + "
  4. KAH115/pT7CFE1-CHis: KAH188/(E/S)/931 + "
  5. KAH148/pT7CFE1-CHis: KAH189/(E/S)/967 + "
  6. KAH150/pT7CFE1-CHis: KAH190/(E/S)/967 + "
  7. KAH151/pT7CFE1-CHis: KAH191/(E/S)/967 + "


Note: KAH109 is being re-done as an internal control to trouble-shoot the ligation
Note: KAH111-KAH151 are all cut and purified already


> Digest (Fermentas FD)

  1. pT7CFE1-CHis, E/P
Reagent Volume  
DNA (plasmid) 25.0 μL
10x buffer 3.0
EcoRI 1.0
PstI 1.0
dH2O 0
  30 μL --> 37°C/ ~30 min.