User:Behzad Damadzadeh/Notebook/PcTF Subcloning in E-coli/2013/01/12 - Revision history

Behzad Damadzadeh: /* 01/12/2013 */

2013-01-12T21:52:14Z

01/12/2013

←Older revision		Revision as of 21:52, 12 January 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.

Behzad Damadzadeh: /* 01/12/2013 */

2013-01-12T21:13:44Z

01/12/2013

←Older revision		Revision as of 21:13, 12 January 2013
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	\|}		\|}

		+	* Calculations are for the ng of insert we need to get a 3:1 ratio of insert molecules to 50 ng vector molecules
	{\| {{table}} cellspacing="3" <!-- Ligation rxn table -->		{\| {{table}} cellspacing="3" <!-- Ligation rxn table -->
	\|   \|\| 1 \|\| 2 \|\| 3 \|\| 4 \|\| 5		\|   \|\| 1 \|\| 2 \|\| 3 \|\| 4 \|\| 5

Behzad Damadzadeh: /* 01/12/2013 */

2013-01-12T21:11:08Z

01/12/2013

←Older revision		Revision as of 21:11, 12 January 2013
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	\|   \|\| 10.52 μL \|\| 10.52 μL \|\| 10μl \|\| 10μL \|\| 10μL		\|   \|\| 10.52 μL \|\| 10.52 μL \|\| 10μl \|\| 10μL \|\| 10μL
	\|}		\|}
		+
		+	* The incubation time for the ligation process was 30min at room temperature.

Behzad Damadzadeh: /* 01/12/2013 */

2013-01-12T21:09:44Z

01/12/2013

←Older revision		Revision as of 21:09, 12 January 2013
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	\| dH<sub>2</sub>O \|\| 0 \|\| 0 \|\| 0.03 \|\| 0.03 \|\| 2.08		\| dH<sub>2</sub>O \|\| 0 \|\| 0 \|\| 0.03 \|\| 0.03 \|\| 2.08
	\|-		\|-
-	\|   \|\| 10.52 μL \|\| 10.52 μL \|\| 10μl \|\| 10μL	+	\|   \|\| 10.52 μL \|\| 10.52 μL \|\| 10μl \|\| 10μL \|\| 10μL
	\|}		\|}

Behzad Damadzadeh: /* 01/12/2013 */

2013-01-12T21:09:24Z

01/12/2013

←Older revision		Revision as of 21:09, 12 January 2013
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	=01/12/2013=		=01/12/2013=
	Ligation of KAH111, KAH112 and C-His vector in DH5α-T and BL21		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 20 μL of nuclease free water, then cut with E/P and purified from the gel.	+	* 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		* Ligations
	{\| {{table}} cellspacing="3" <!-- Ligations table -->		{\| {{table}} cellspacing="3" <!-- Ligations table -->
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	{\| {{table}} cellspacing="3" <!-- Ligation rxn table -->		{\| {{table}} cellspacing="3" <!-- Ligation rxn table -->
-	\|   \|\| 1 \|\| 2 \|\| 3 \|\|	+	\|   \|\| 1 \|\| 2 \|\| 3 \|\| 4 \|\| 5
	\|-		\|-
-	\| Insert DNA \|\| 3.12 \|\| 3.12 \|\| --- \|\|	+	\| Insert DNA \|\| 2.34 \|\| 2.34 \|\| 2.05 \|\| 2.05 \|\| ---
	\|-		\|-
-	\| Vector DNA \|\| 2.94 \|\| 2.94 \|\| 2.94 \|\|	+	\| Vector DNA \|\| 1.92 \|\| 1.92 \|\| 1.92 \|\| 1.92 \|\| 1.92
	\|-		\|-
-	\| 2x lgn buf (Roche) \|\| 7.06 \|\| 7.06 \|\| 5.0 \|\|	+	\| 2x lgn buf (Roche) \|\| 5.26 \|\| 5.26 \|\| 5.0 \|\| 5.0 \|\| 5.0
	\|-		\|-
-	\| T4 ligase (NEB) \|\| 1.0 \|\| 1.0 \|\| 1.0 \|\|	+	\| T4 ligase (NEB) \|\| 1.0 \|\| 1.0 \|\| 1.0 \|\| 1.0 \|\| 1.0
	\|-		\|-
-	\| dH<sub>2</sub>O \|\| 0 \|\| 0 \|\| 0.88 \|\|	+	\| dH<sub>2</sub>O \|\| 0 \|\| 0 \|\| 0.03 \|\| 0.03 \|\| 2.08
	\|-		\|-
-	\|   \|\| 14.12 μL \|\| 14.12 μL \|\| 10μl \|\|	+	\|   \|\| 10.52 μL \|\| 10.52 μL \|\| 10μl \|\| 10μL
	\|}		\|}

Behzad Damadzadeh: /* 01/12/2013 */

2013-01-12T21:05:18Z

01/12/2013

Behzad Damadzadeh: /* 01/12/2013 */

2013-01-12T21:00:13Z

01/12/2013

←Older revision		Revision as of 21:00, 12 January 2013
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	{\| {{table}} cellspacing="3" <!-- Ligations table -->		{\| {{table}} cellspacing="3" <!-- Ligations table -->
	\|- bgcolor=#cfcfcf		\|- bgcolor=#cfcfcf
-	\| Ligation \|\| <font color="blue"><u>~~Plate results (lig : neg crtl)</u> 12/12/12~~</font>	+	\| Ligation \|\| <font color="blue"><u></font>
	\|-		\|-
	\| 1. E. coli DH5α-T KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>		\| 1. E. coli DH5α-T KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>
	\|-		\|-
-	\| 2. E. coli BL-21 KAH111(E/P)/size, 16 ng + C-His(E/P)/3600, ~~25ng~~ \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>	+	\| 2. E. coli BL-21 KAH111(E/P)/size, 16 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>
		+	\|-
		+	\| 1. E. coli DH5α-T KAH112(E/P)/size, 22.8 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH112/C-His 3:1 No Colonies</font>
		+	\|-
		+	\| 2. E. coli BL-21 KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH112/C-His 3:1 No Colonies</font>
	\|-		\|-
	\| 3. C-His(E/P)/ 25 ng \|\|		\| 3. C-His(E/P)/ 25 ng \|\|

Behzad Damadzadeh: /* Date */

2013-01-12T20:57:54Z

Date

←Older revision		Revision as of 20:57, 12 January 2013
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	<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->		<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->

-	=~~Date~~=	+	=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 20 μL of nuclease free water, then cut with E/P and purified from the gel.
		+	* Ligations
		+	{\| {{table}} cellspacing="3" <!-- Ligations table -->
		+	\|- bgcolor=#cfcfcf
		+	\| Ligation \|\| <font color="blue"><u>Plate results (lig : neg crtl)</u> 12/12/12</font>
		+	\|-
		+	\| 1. E. coli DH5α-T KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>
		+	\|-
		+	\| 2. E. coli BL-21 KAH111(E/P)/size, 16 ng + C-His(E/P)/3600, 25ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>
		+	\|-
		+	\| 3. C-His(E/P)/ 25 ng \|\|
		+	\|}

-	~~'''List title'''~~	+	{\| {{table}} cellspacing="3" <!-- Ligation rxn table -->
-	~~# List items~~	+	\|   \|\| 1 \|\| 2 \|\| 3
		+	\|-
		+	\| Insert DNA \|\| 3.12 \|\| 3.12 \|\| ---
		+	\|-
		+	\| Vector DNA \|\| 2.94 \|\| 2.94 \|\| 2.94
		+	\|-
		+	\| 2x lgn buf (Roche) \|\| 7.06 \|\| 7.06 \|\| 5.0
		+	\|-
		+	\| T4 ligase (NEB) \|\| 1.0 \|\| 1.0 \|\| 1.0
		+	\|-
		+	\| dH<sub>2</sub>O \|\| 0 \|\| 0 \|\| 0.88
		+	\|-
		+	\|   \|\| 14.12 μL \|\| 14.12 μL \|\| 10μl
		+	\|}

Behzad Damadzadeh: Autocreate 2013/01/12 Entry for User:Behzad_Damadzadeh/Notebook/PcTF_Subcloning_in_E-coli

2013-01-12T20:51:17Z

Autocreate 2013/01/12 Entry for User:Behzad_Damadzadeh/Notebook/PcTF_Subcloning_in_E-coli

New page

{|{{table}} width="800"
|-
|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">PcTF Subcloning in E. coli</span>
|style="background-color: #F2F2F2" align="center"|<html><img src="/images/9/94/Report.png" border="0" /></html> [[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>[[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]]<html>      </html>}}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]]<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>}}
|-
| colspan="2"|


=Date=

'''List title'''
# List items

←Older revision		Revision as of 21:05, 12 January 2013
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	\| Ligation \|\| <font color="blue"><u></font>		\| Ligation \|\| <font color="blue"><u></font>
	\|-		\|-
-	\| 1. E. coli ~~DH5α~~-T KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>	+	\| 1. E. coli BL-21 KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>
	\|-		\|-
-	\| 2. E. coli BL-21 KAH111(E/P)/size, 16 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>	+	\| 2. E. coli DH5α-T KAH111(E/P)/size, 16 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH111/C-His 3:1 No Colonies</font>
	\|-		\|-
-	\| 1. E. coli ~~DH5α~~-T KAH112(E/P)/size, 22.8 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH112/C-His 3:1 No Colonies</font>	+	\| 3. E. coli BL-21 KAH112(E/P)/size, 22.8 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH112/C-His 3:1 No Colonies</font>
	\|-		\|-
-	\| 2. E. coli BL-21 KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH112/C-His 3:1 No Colonies</font>	+	\| 4. E. coli DH5α-T KAH111(E/P)/size, 20 ng + C-His(E/P)/3600, 26ng \|\| <font color="blue">KAH112/C-His 3:1 No Colonies</font>
	\|-		\|-
-	\| 3. C-His(E/P)/ 25 ng \|\|	+	\| 5. C-His(E/P)/ 26 ng (Control Plate)\|\|
	\|}		\|}

	{\| {{table}} cellspacing="3" <!-- Ligation rxn table -->		{\| {{table}} cellspacing="3" <!-- Ligation rxn table -->
-	\|   \|\| 1 \|\| 2 \|\| 3	+	\|   \|\| 1 \|\| 2 \|\| 3 \|\|
	\|-		\|-
-	\| Insert DNA \|\| 3.12 \|\| 3.12 \|\| ---	+	\| Insert DNA \|\| 3.12 \|\| 3.12 \|\| --- \|\|
	\|-		\|-
-	\| Vector DNA \|\| 2.94 \|\| 2.94 \|\| 2.94	+	\| Vector DNA \|\| 2.94 \|\| 2.94 \|\| 2.94 \|\|
	\|-		\|-
-	\| 2x lgn buf (Roche) \|\| 7.06 \|\| 7.06 \|\| 5.0	+	\| 2x lgn buf (Roche) \|\| 7.06 \|\| 7.06 \|\| 5.0 \|\|
	\|-		\|-
-	\| T4 ligase (NEB) \|\| 1.0 \|\| 1.0 \|\| 1.0	+	\| T4 ligase (NEB) \|\| 1.0 \|\| 1.0 \|\| 1.0 \|\|
	\|-		\|-
-	\| dH<sub>2</sub>O \|\| 0 \|\| 0 \|\| 0.88	+	\| dH<sub>2</sub>O \|\| 0 \|\| 0 \|\| 0.88 \|\|
	\|-		\|-
-	\|   \|\| 14.12 μL \|\| 14.12 μL \|\| 10μl	+	\|   \|\| 14.12 μL \|\| 14.12 μL \|\| 10μl \|\|
	\|}		\|}