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Team:Bielefeld-Germany/Labjournal/Molecular
From 2013.igem.org
(Difference between revisions)
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<tr><th colspan="5">Program Phusion</th></tr> | <tr><th colspan="5">Program Phusion</th></tr> | ||
<tr align="center"><td>98 °C</td><td>98 °C</td><td>60 °C</td><td>72 °C</td><td>72 °C</td></tr> | <tr align="center"><td>98 °C</td><td>98 °C</td><td>60 °C</td><td>72 °C</td><td>72 °C</td></tr> | ||
| - | <tr align="center"><td>1:00</td> <td> | + | <tr align="center"><td>1:00</td> <td>10 sec</td><td>30 sec</td> <td>30 sec</td><td>6:00</td></tr> |
<tr><td></td><td colspan="3" align="center">35 cycles</td><td></td></tr> | <tr><td></td><td colspan="3" align="center">35 cycles</td><td></td></tr> | ||
<tr><td colspan="5"> | <tr><td colspan="5"> | ||
Revision as of 20:16, 30 August 2013
PCR Protocols and Programs
Phusion 50ul for mtrCAB Fragment1
| 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 -> 22sec/kb | ||||
Phusion 50ul for mtrCAB Fragment2
| 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 | 60 °C | 72 °C | 72 °C |
| 1:00 | 10 sec | 30 sec | 30 sec | 6:00 |
| 35 cycles | ||||
|
Notes:
Size: 330bp -> 90sec/kb | ||||
Taq 25 µl
| Reagenz | 1x | 8x |
|---|---|---|
| Buffer 5x | 5,0 | 44 |
| MgSO4 | 0 | 0 |
| dNTPs | 2,5 | 22 |
| Primer1 | 1,0 | 8,8 |
| Primer2 | 1,0 | 8,8 |
| Template | 1,5 | 13,2 |
| DMSO | 2,0 | 17,6 |
| Polymerase | 0,5 | 4,4 |
| H2O | 11,5 | 101,2 |
| Program Taq | ||||
|---|---|---|---|---|
| 98 °C | 98 °C | x °C | 72 °C | 72 °C |
| 2 min | 30 s | 30 s | y | 6 min |
| x: according to Primer Tm y: 15s/kb | ||||
