18S- S30/5FK 20101011

Procedure

PCR cocktail: 
Component Volume (ul) Mix (ul)
Forward primer  0.5  
Reversed primer  0.5  
Buffer    
MgCl2    
dNTPs    
Taq    
DNA (if nested)  1  
ddH2O
 23  
Total volume  25
Forward primer(s): 
Thermal cycling (abbreviated): 
95ºC@4min, 35X(94ºC@30sec,52ºC@30sec, 72ºC@30sec)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
240
95  
Denaturation DNA melting producing single-stranded DNA      
Amplification   35 -  -
Denaturation DNA melting producing single-stranded DNA
 - 30 94 
Annealing The primer binds to the template  - 30  52
Extension The primer binds to the template
 - 30  72
Final elongation Any remaining single-stranded DNA is extended 1 600  72
Hold   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