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Team:Newcastle/PCR
From 2010.igem.org
(Difference between revisions)
(New page: {{Team:Newcastle/mainbanner}} ==PCR== ====Materials to add accordingly:==== # 37.5 µl of distilled H2O # 10 µl of 5x GoTaq Buffer # Nucleotide DNTP # 2.5 µl forward primer # 2.5 µl ...) |
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{{Team:Newcastle/mainbanner}} | {{Team:Newcastle/mainbanner}} | ||
| - | ==PCR== | + | ==GoTaq PCR== |
| - | + | ===Materials to add accordingly:=== | |
# 37.5 µl of distilled H2O | # 37.5 µl of distilled H2O | ||
# 10 µl of 5x GoTaq Buffer | # 10 µl of 5x GoTaq Buffer | ||
| - | # | + | # 1 µl of nucleotide DNTP |
# 2.5 µl forward primer | # 2.5 µl forward primer | ||
# 2.5 µl backward primer | # 2.5 µl backward primer | ||
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| - | + | ===Conditions for ThermoCycler:=== | |
# Initialise - 95°C for 2 minutes. | # Initialise - 95°C for 2 minutes. | ||
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Steps 2 to 4 are repeated for 30 cycles before continuing to step 5. | Steps 2 to 4 are repeated for 30 cycles before continuing to step 5. | ||
| + | |||
| + | ==Phusion PCR== | ||
| + | |||
| + | ===Materials to add accordingly:=== | ||
| + | |||
| + | # 27.5 µl of distilled H2O | ||
| + | # 10 µl of 5x Buffer | ||
| + | # 1 µl of nucleotide DNTP | ||
| + | # 5 µl forward primer | ||
| + | # 5 µl backward primer | ||
| + | # 1 µl template DNA | ||
| + | #0.5 µl of Fusion | ||
{{Team:Newcastle/footer}} | {{Team:Newcastle/footer}} | ||
Revision as of 14:00, 27 July 2010
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Contents |
GoTaq PCR
Materials to add accordingly:
- 37.5 µl of distilled H2O
- 10 µl of 5x GoTaq Buffer
- 1 µl of nucleotide DNTP
- 2.5 µl forward primer
- 2.5 µl backward primer
- 1 µl template DNA
Conditions for ThermoCycler:
- Initialise - 95°C for 2 minutes.
- Denature - 95°C for 30 seconds.
- Anneal - 52°C for 30 seconds (melting temperature, Tm, of template)
- Extension - 75°C for 30 seconds
- Extension finish - 75°C for 5 minutes
- Hold - 4°C
Steps 2 to 4 are repeated for 30 cycles before continuing to step 5.
Phusion PCR
Materials to add accordingly:
- 27.5 µl of distilled H2O
- 10 µl of 5x Buffer
- 1 µl of nucleotide DNTP
- 5 µl forward primer
- 5 µl backward primer
- 1 µl template DNA
- 0.5 µl of Fusion
   
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