28S-UJ2176/L3449 20091203

Basic information

Procedure

PCR cocktail: 
Component Volume (ul) Mix (ul)
Forward primer  0,5  
Reversed primer  0,5  
Buffer    
MgCl2    
dNTPs    
Taq    
DNA (if nested)  2  
ddH2O
 22  
Total volume  25
Forward primer(s): 
Thermal cycling (abbreviated): 
95ºC@4min, 2x(94ºC@30sec, 58-54ºC@30sec,72ºC@60), 34x(94ºC@30sec, 52ºC@30sec, 72ºC@60sec),72ºC@10min
PCR machine: 
GeneAmp
Thermal cycling: 
Step Comment Cycles Time (sec) Temperature (°C)
Initialization For DNA polymerases that need hot-start heat activation 1
 
Denaturation DNA melting producing single-stranded DNA 1 240 95 
Amplification   2 -  -
Denaturation DNA melting producing single-stranded DNA
 - 30  94
Annealing The primer binds to the template  - 30  58-54
Extension The DNA polymerase synthesizes new DNA  - 60  72
Amplification   34    
Denaturation    30 94
Annealing   30 52
Extension   60 72
Final elongation Any remaining single-stranded DNA is extended   600 72
Final hold Short-term storage 1 -  12
Reversed primer(s): 
File attachments: 
Products and Equipment: 

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Scratchpads developed and conceived by (alphabetical): Ed Baker, Katherine Bouton Alice Heaton Dimitris Koureas, Laurence Livermore, Dave Roberts, Simon Rycroft, Ben Scott, Vince Smith