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Team:Bielefeld-Germany/Labjournal/PCRProtocols
From 2013.igem.org
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<h1 style="color:#ff6600;">PCR Protocols and Programs</h1> | <h1 style="color:#ff6600;">PCR Protocols and Programs</h1> | ||
<h2>Standard Protocols</h2> | <h2>Standard Protocols</h2> | ||
Revision as of 22:02, 25 September 2013
PCR Protocols and Programs
Standard Protocols
| Reagent | 1x | 8x |
|---|---|---|
| Hf-Buffer 5x | 10.0 | 88.0 |
| dNTPs (10mM) | 2.0 | 17.6 |
| Primer1 (10mM) | 1.0 | 8.8 |
| Primer2 (10mM) | 1.0 | 8.8 |
| Template (16.5ng) | 3.0 | 26.4 |
| DMSO | 4.0 | 35.2 |
| Phusion (0.5u) | 1.0 | 8.8 |
| H2O | 26.0 | 228.8 |
Specific Protocols
| Reagent | 1x | 8x |
|---|---|---|
| Hf-Buffer 5x | 10.0 | 88.0 |
| dNTPs (10mM) | 2.0 | 17.6 |
| Primer1 (10mM) | 1.0 | 8.8 |
| Primer2 (10mM) | 1.0 | 8.8 |
| Template (16.5ng) | 3.0 | 26.4 |
| DMSO | 4.0 | 35.2 |
| Phusion (0.5u) | 1.0 | 8.8 |
| H2O | 26.0 | 228.8 |
| Program Phusion | ||||
|---|---|---|---|---|
| 98 °C | 98 °C | 68 °C | 72 °C | 72 °C |
| 30s | 10s | 30s | 1:35 | 6:00 |
| 35 cycles | ||||
|
Notes:
Size: 1800bp Primer1: Primer2: | ||||
| Reagent | 1x | 8x |
|---|---|---|
| Hf-Buffer 5x | 10.0 | 88.0 |
| dNTPs (10mM) | 2.5 | 22.0 |
| Primer1 (10mM) | 1.0 | 8.8 |
| Primer2 (10mM) | 1.0 | 8.8 |
| Template (5.5ng/ul) | 2.0 | 17.6 |
| DMSO | 3.0 | 26.4 |
| Phusion (0.5u) | 1.0 | 8.8 |
| H2O | 29.5 | 259.6 |
| Program Phusion | ||||
|---|---|---|---|---|
| 98 °C | 98 °C | 60 °C | 72 °C | 72 °C |
| 1:00 | 10s | 30s | 30s | 6:00 |
| 35 cycles | ||||
|
Notes:
Size: 330bp Primer1: Primer2: | ||||
| Reagent | 1x | 8x |
|---|---|---|
| Hf-Buffer 5x | 10.0 | 88.0 |
| dNTPs (10mM) | 2.5 | 22.0 |
| Primer1 (10mM) | 1.0 | 8.8 |
| Primer2 (10mM) | 1.0 | 8.8 |
| Template (5.5ng/ul) | 2.0 | 17.6 |
| DMSO | 3.0 | 26.4 |
| Phusion (0.5u) | 1.0 | 8.8 |
| H2O | 29.5 | 259.6 |
| Program Phusion | ||||
|---|---|---|---|---|
| 98 °C | 98 °C | 70 °C | 72 °C | 72 °C |
| 30s | 10s | 30s | 50s | 6:00 |
| 35 cycles | ||||
|
Notes:
Size: 3000bp Primer1: Primer2: | ||||
| Reagent | 1x | 8x |
|---|---|---|
| GC-Buffer 5x | 10.0 | 88.0 |
| dNTPs (10mM) | 2.0 | 17.6 |
| Primer1 (10mM) | 1.25 | 11.0 |
| Primer2 (10mM) | 1.25 | 11.0 |
| Template (5.5ng/ul) | 1.0 | 8.8 |
| DMSO | 1.5 | 13.2 |
| Phusion (0.5u) | 0.5 | 4.4 |
| H2O | 32.5 | 286.0 |
| Program Phusion | ||||
|---|---|---|---|---|
| 98 °C | 98 °C | 70 °C | 72 °C | 72 °C |
| 45s | 30s | 1:00 | 45s | 5:00 |
| 35 cycles | ||||
|
Notes:
Size: 2070bp Primer1_Gibson: Primer2_Gibson: Primer1: Primer2: | ||||
| Reagent | 1x | 8x |
|---|---|---|
| Hf-Buffer 5x | 10.0 | 88.0 |
| dNTPs (10mM) | 2.5 | 22.0 |
| Primer1 (10mM) | 1.0 | 8.8 |
| Primer2 (10mM) | 1.0 | 8.8 |
| Template (5.5ng/ul) | 2.0 | 17.6 |
| DMSO | 3.0 | 26.4 |
| Phusion (0.5u) | 1.0 | 8.8 |
| H2O | 29.5 | 259.6 |
| Program Phusion | ||||
|---|---|---|---|---|
| 98 °C | 98 °C | 60 °C | 72 °C | 72 °C |
| 30s | 10s | 30s | 1:15 | 6:15 |
| 35 cycles | ||||
|
Notes:
Size: 5150bp Primer1: Primer2: | ||||
| Reagent | 1x | 8x |
|---|---|---|
| Hf-Buffer 5x | 10.0 | 88.0 |
| dNTPs (10mM) | 2.0 | 17.6 |
| Primer1 (10mM) | 1.0 | 8.8 |
| Primer2 (10mM) | 1.0 | 8.8 |
| Template (5.5ng/ul) | 3.0 | 26.4 |
| DMSO | 3.0 | 26.4 |
| Phusion (0.5u) | 1.0 | 8.8 |
| H2O | 29.5 | 255.2 |
| Program Phusion | ||||
|---|---|---|---|---|
| 98 °C | 98 °C | 68 °C | 72 °C | 72 °C |
| 30s | 10s | 30s | 1:35 | 6:00 |
| 35 cycles | ||||
|
Notes:
Size: 6311bp Primer1: Primer2: | ||||
