Go Back   Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum > Molecular Research Topics Forum > PCR - Polymerase Chain Reaction Forum
No amplification using pfu polymerase No amplification using pfu polymerase
Register Search Today's Posts Mark Forums Read

PCR - Polymerase Chain Reaction Forum PCR - Polymerase Chain Reaction Forum. Discuss and ask questions about PCR troubleshooting, PCR protocols and methods, PCR products, and PCR theory.


No amplification using pfu polymerase

No amplification using pfu polymerase - PCR - Polymerase Chain Reaction Forum

No amplification using pfu polymerase - PCR - Polymerase Chain Reaction Forum. Discuss and ask questions about PCR troubleshooting, PCR protocols and methods, PCR products, and PCR theory.


Reply
 
LinkBack Thread Tools Display Modes
  #1  
Old 04-01-2007, 09:18 AM
Pipette Filler
Points: 1,350, Level: 21 Points: 1,350, Level: 21 Points: 1,350, Level: 21
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Apr 2007
Posts: 1
Thanks: 0
Thanked 0 Times in 0 Posts
Question No amplification using pfu polymerase


Urgent help needed....
I am trying to amplify a gene from strep chormosomal DNA extraction to clone. Works beautifully with TAQ but when i try the same conditions using pfu hot start doesn't work at all.............Have tried manufacturers instructions..doesn't work!! getting desperate...Please trouble shoot...

Strep girl
Reply With Quote
  #2  
Old 04-25-2007, 05:39 PM
Pipette Filler
Points: 1,273, Level: 20 Points: 1,273, Level: 20 Points: 1,273, Level: 20
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Apr 2007
Posts: 1
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: No amplification using pfu polymerase
Have you noticed that pfu polymerase has lower extension rate?
Reply With Quote
  #3  
Old 05-24-2007, 10:10 AM
Pipette Filler
Points: 1,294, Level: 20 Points: 1,294, Level: 20 Points: 1,294, Level: 20
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: May 2007
Posts: 7
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: No amplification using pfu polymerase
I find it can help to try a range of MgCl2 concentrations in your PCR mix. You'd be suprised what an impact it can have.
Reply With Quote
  #4  
Old 05-28-2007, 09:48 AM
Pipette Filler
Points: 1,594, Level: 23 Points: 1,594, Level: 23 Points: 1,594, Level: 23
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Nov 2006
Posts: 2
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: No amplification using pfu polymerase
It require very high qulity of your template DNA. Did you check your DNA qulity?
Reply With Quote
  #5  
Old 06-11-2007, 04:01 PM
Pipette Filler
Points: 2,042, Level: 28 Points: 2,042, Level: 28 Points: 2,042, Level: 28
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: May 2007
Posts: 11
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: No amplification using pfu polymerase
I had the same problem with pfu but by using a fresh DNA it worked very well.
try to purify ur DNA sample or fresh extract the dna.
Reply With Quote
  #6  
Old 07-10-2007, 11:27 AM
ale ale is offline
Pipette Filler
Points: 1,143, Level: 19 Points: 1,143, Level: 19 Points: 1,143, Level: 19
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Jul 2007
Posts: 2
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: No amplification using pfu polymerase
i am really sorry to disturb you but i would like to tell you that right now i am working on an amplification of a gene with pfu and unfortunately it does not work so i was wondering what have you done and if you have resolved the problem. please let me know as soon as possible because these experiments are really important for my university thesis
Reply With Quote
  #7  
Old 07-16-2007, 02:01 AM
Pipette Filler
Points: 2,042, Level: 28 Points: 2,042, Level: 28 Points: 2,042, Level: 28
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: May 2007
Posts: 11
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: No amplification using pfu polymerase
hello

you should fresh DNA sample.IT works very well when fresh DNA sample is used.
You should also use a blend of Taq nad Pfu.I use 1:2 ratio of taq and pfu.
Reply With Quote
Reply

Tags
amplification , pfu , polymerase


« PFU PCR Amplification Problems | First PCR Worked, Second PCR Did not Work »
Thread Tools
Display Modes
Linear Mode Linear Mode

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts
BB code is On
Smilies are On
[IMG] code is On
HTML code is Off
Trackbacks are On
Pingbacks are On
Refbacks are On
Forum Jump

Similar Threads
Thread Thread Starter Forum Replies Last Post
Weird amplification curve for Taqman probes fen Real-Time PCR and Quantitative PCR Forum 0 01-25-2010 08:35 AM
The polymerase chain reaction (PCR) requires __________ and __________ to be Receive PCR - Polymerase Chain Reaction Forum 7 10-21-2009 06:28 AM
PCR World-Everything about Polymerase Chain Reaction peterish PCR - Polymerase Chain Reaction Forum 0 11-27-2008 08:12 PM
Pfu polymerase mixed with Taq polymerase Julio Cortazar Microbiology Forum 6 05-29-2006 08:33 AM
patents/licenses and Taq/Pfu DNA polymerase news.uni-muenster.de Protocols and Methods Forum 6 05-06-2005 04:13 PM


All times are GMT. The time now is 01:23 PM.

Contact Us - Molecular Biology - Archive - Privacy Statement - Top

Powered by vBulletin® Version 3.8.4
Copyright ©2000 - 2013, Jelsoft Enterprises Ltd.
Copyright 2005 - 2012 Molecular Station | All Rights Reserved
Page generated in 0.18210 seconds with 16 queries