Team:SDU-Denmark/labwork/Protocols/PCR

From 2012.igem.org

(Difference between revisions)
Line 287: Line 287:
72°C for 5-10 minutes </br>
72°C for 5-10 minutes </br>
4°C on hold</pre>
4°C on hold</pre>
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</br>
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</br>
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<h2>Non-proof-reading PCR using Taq Polymerase</h2>
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 +
5μl Dream Taq Buffer </br>
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5μl dNTP</br>
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1μl VF2</br>
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1μl VR</br>
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Udtag 10μl fra primer stock og fortynd i 90μl oprenset vand</br>
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1,5μl Dream Taq Polymerase</br>
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1μl Template</br>
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H2O up to 50μl </br>
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<b>PCR-PROGRAM:</b>
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1. Start 95°C 2min </br>
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2. Denaturing 95°C 1min </br>
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3. Annealing 55°C 30sek</br>
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4. Elonging 72°C 1min (1kb/min) </br>
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5. GOTO 2, x 29 cycles</br>
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6. End 72°C 5min</br>
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7. Hold 4°C infinite </br>
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Revision as of 13:36, 12 September 2012

iGEM TEAM ::: SDU-DENMARK courtesy of NIAID

mRNA Isolation PCR Miniprep Check Digest
Content Content Content Content
Content Content Content Content

Polymerase Chain Reaction

Proof-reading PCR using Phusion Hot Start II

some_text

PCR program: 
95°C for 30 seconds 

25-35 cycles:
98°C for 30 seconds 
Annealing temperature (5°C below primer melting temperature)  
72°C for 15-30s/kb

72°C for 5-10 minutes 

4°C on hold


Non-proof-reading PCR using Taq Polymerase

5μl Dream Taq Buffer
5μl dNTP
1μl VF2
1μl VR
Udtag 10μl fra primer stock og fortynd i 90μl oprenset vand
1,5μl Dream Taq Polymerase
1μl Template
H2O up to 50μl
PCR-PROGRAM: 1. Start 95°C 2min
2. Denaturing 95°C 1min
3. Annealing 55°C 30sek
4. Elonging 72°C 1min (1kb/min)
5. GOTO 2, x 29 cycles
6. End 72°C 5min
7. Hold 4°C infinite

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