Team:EPF-Lausanne/Protocols/1.5 steps PCR

From 2009.igem.org

(Difference between revisions)

Latest revision as of 18:44, 20 October 2009

1.5 Steps PCR Protocol

1. Do the following mixing:

dNTP	1 μl
5x Buffer + MgCl2	10 μl
Primer Mix	1 μl
template	0.5 μl
HiFi Plus DNAp	0.5 μl
dH2O	36.5 μl
Final Volume	50 μl

2. Make it run 10 cycles:

1	94°C	4:00
2	94°C	0:30
3	55°C	1:00
4	72°C	2:00
5	Cycle 2-4 10 times
6	72°C	7:00
7	4°C	∞

3. We now add the standard iGEM primers which finish the extension, and do the following cycles:

1	94°C	4:00
2	94°C	0:30
3	55°C	1:00
4	72°C	2:00
5	Cycle 2-4 25-30 times
6	72°C	7:00
7	4°C	∞

@@ Line 15: / Line 15: @@
 <html><center>
-<font size="6" color="#007CBC"><i>Miniprep Protocol</i></font>
+<font size="6" color="#007CBC">1.5 Steps PCR Protocol</font>
 </center></html>
-<br>
 <br>
 ----
@@ Line 56: / Line 55: @@
 {| class="wikitable" style="text-align:left; width:20%;"
 |-
-! scope=row | 1
+! scope=row | 1&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
 |94°C
 |4:00
 |-
-! scope=row | 2
+! scope=row | 2&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
 |94°C
 |0:30
 |-
-! scope=row | 3
+! scope=row | 3&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
 |55°C
 |1:00
 |-
-! scope=row | 4
+! scope=row | 4&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
 |72°C
 |2:00
 |-
-! scope=row | 5
+! scope=row | 5&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
 |Cycle 2-4 10 times
 |
 |-
-! scope=row | 6
+! scope=row | 6&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
+|72°C
+|7:00
+|-
+! scope=row | 7&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
+|4°C
+|&infin;
+|}
+</center>
+'''3.''' We now add the standard iGEM primers which finish the extension, and do the following cycles:
+<center>
+{| class="wikitable" style="text-align:left; width:20%;"
+|-
+! scope=row | 1&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
+|94°C
+|4:00
+|-
+! scope=row | 2&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
+|94°C
+|0:30
+|-
+! scope=row | 3&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
+|55°C
+|1:00
+|-
+! scope=row | 4&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
+|72°C
+|2:00
+|-
+! scope=row | 5&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
+|Cycle 2-4 25-30 times
+|
+|-
+! scope=row | 6&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
 |72°C
 |7:00
 |-
-! scope=row | 7
+! scope=row | 7&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;
 |4°C
 |&infin;