Team:Heidelberg/Templates/DelH week24

From 2013.igem.org

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== 23-09 - 29-09-13 ==
== 23-09 - 29-09-13 ==
===Generation of DelH plasmids pFS02, pFS03, pFS04 and pFS05===
===Generation of DelH plasmids pFS02, pFS03, pFS04 and pFS05===
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{| class="wikitable"
{| class="wikitable"
|-
|-
-
! Reagent !! Amount   
+
! Reagent !! Amount [µl]    
|-
|-
| pSB6A1 || 1
| pSB6A1 || 1
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{| class="wikitable"
{| class="wikitable"
|-
|-
-
! Reagent !! Amount  
+
! Reagent !! Amount [µl] 
|-
|-
| pSB6A1 || 1
| pSB6A1 || 1
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{| class="wikitable"
{| class="wikitable"
|-
|-
-
! Reagent !! Amount   
+
! Reagent !! Amount [µl]    
|-
|-
| pSB6A1 || 1
| pSB6A1 || 1
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{| class="wikitable"
{| class="wikitable"
|-
|-
-
! Reagent !! Amount  
+
! Reagent !! Amount [µl] 
|-
|-
| pSB4K5 || 1
| pSB4K5 || 1
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'''Results:'''
'''Results:'''
-
====Amplification of pFS_02 Backbone (FS_86 to FS_87; 4.1 kbp)====
+
====Amplification of DelH fragment (FS_84 to FS_66; 4.1 kbp)====
:'''Reaction'''  
:'''Reaction'''  
{| class="wikitable"
{| class="wikitable"
|-
|-
-
! Reagent !! Amount   
+
! Reagent !! Amount [µl]    
|-
|-
-
| pSB4K5 || 1
+
| ''D. acidovorans'' SPH-1 || 1
|-
|-
| FS_86:  (1/10) || 1
| FS_86:  (1/10) || 1
|-
|-
| FS_87:  (1/10) || 1
| FS_87:  (1/10) || 1
 +
|-
 +
| Phusion flash Master Mix || 10
 +
|-
 +
| DMSO || 1
 +
|-
 +
| dd H<sub>2</sub>O || 4
 +
|}
 +
 +
:'''Conditions '''
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 +
{| class="wikitable"
 +
|-
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! colspan="3" | Biorad T100
 +
|-
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! Cycles !! temperature [°C] !! Time [s]
 +
|-
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| 1 || 98 || 10
 +
|-
 +
| rowspan="2"| 35 || 98 || 1
 +
|-
 +
| 72  || 1:25
 +
|-
 +
| 1 || 72 || 10min
 +
|-
 +
| 1  || 12 || inf
 +
|}
 +
 +
'''Results:'''
 +
 +
====Amplification of ccdB fragment for pFS_04 (FS_90 to FS_93; 700 bp)====
 +
:'''Reaction'''
 +
{| class="wikitable"
 +
|-
 +
! Reagent !! Amount [µl] 
 +
|-
 +
| pDonor-Plasmid || 1
 +
|-
 +
| FS_90:  (1/10) || 1
 +
|-
 +
| FS_93:  (1/10) || 1
|-
|-
| Phusion flash Master Mix || 10
| Phusion flash Master Mix || 10

Latest revision as of 15:35, 22 October 2013

Contents

23-09 - 29-09-13

Generation of DelH plasmids pFS02, pFS03, pFS04 and pFS05

Amplification of pFS_04 Backbone (FS_85 to FS_94; 4.1 kbp)

Reaction
Reagent Amount [µl]
pSB6A1 1
FS_85: (1/10) 1
FS_94: (1/10) 1
Phusion flash Master Mix 10
DMSO 1
dd H2O 4
Conditions
Biorad T100
Cycles temperature [°C] Time [s]
1 98 10
35 98 1
72 1:25
1 72 10min
1 12 inf

Results:

Amplification of pFS_02 Backbone (HM_17 to FS_78; 4.1 kbp)

Reaction
Reagent Amount [µl]
pSB6A1 1
HM_17: (1/10) 1
FS_78: (1/10) 1
Phusion flash Master Mix 10
DMSO 1
dd H2O 4
Conditions
Biorad T100
Cycles temperature [°C] Time [s]
1 98 10
35 98 1
72 1:25
1 72 10min
1 12 inf

Results:

Amplification of pFS_02 Backbone (HM_17 to FS_85; 4.1 kbp)

Reaction
Reagent Amount [µl]
pSB6A1 1
HM_17: (1/10) 1
FS_85: (1/10) 1
Phusion flash Master Mix 10
DMSO 1
dd H2O 4
Conditions
Biorad T100
Cycles temperature [°C] Time [s]
1 98 10
35 98 1
72 1:25
1 72 10min
1 12 inf

Results:

Amplification of pFS_03 Backbone (FS_86 to FS_87; 4.1 kbp)

Reaction
Reagent Amount [µl]
pSB4K5 1
FS_86: (1/10) 1
FS_87: (1/10) 1
Phusion flash Master Mix 10
DMSO 1
dd H2O 4
Conditions
Biorad T100
Cycles temperature [°C] Time [s]
1 98 10
35 98 1
72 1:25
1 72 10min
1 12 inf

Results:

Amplification of DelH fragment (FS_84 to FS_66; 4.1 kbp)

Reaction
Reagent Amount [µl]
D. acidovorans SPH-1 1
FS_86: (1/10) 1
FS_87: (1/10) 1
Phusion flash Master Mix 10
DMSO 1
dd H2O 4
Conditions
Biorad T100
Cycles temperature [°C] Time [s]
1 98 10
35 98 1
72 1:25
1 72 10min
1 12 inf

Results:

Amplification of ccdB fragment for pFS_04 (FS_90 to FS_93; 700 bp)

Reaction
Reagent Amount [µl]
pDonor-Plasmid 1
FS_90: (1/10) 1
FS_93: (1/10) 1
Phusion flash Master Mix 10
DMSO 1
dd H2O 4
Conditions
Biorad T100
Cycles temperature [°C] Time [s]
1 98 10
35 98 1
72 1:25
1 72 10min
1 12 inf

Results:

Retrieved from "http://2013.igem.org/Team:Heidelberg/Templates/DelH_week24"