PCR questions?

Duncan Clark Duncan at genesys.demon.co.uk
Wed Aug 17 10:20:05 EST 1994

In article: <01HFXWRP2NN695MW1L at SKYCAT.USask.CA>  LEES at SASK.USASK.CA writes:

> 1. What exactly does final extension-used be 10 min after last 
> cycle-work for?

This in theory gives the polymerase time to fill in  any ragged ends ie
blunts the amplification product but see next answer.

> 2. Why does PCR product have "A" overhang?

Most non-proofreading polymerases have an associated terminal 
transferase activity which in Taq's case prefers to stick an 
'A' on the end. It can however be forced to use other bases. The 10 min
final extension also helps to put the 'A' on the end.

Duncan Clark                        | Internet:    duncan at genesys.demon.co.uk
G4ELJ                               | Compuserve:  100015.1406 at compuserve.com

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