Go Back   Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum > Molecular Research Topics Forum > PCR - Polymerase Chain Reaction Forum > Real-Time PCR and Quantitative PCR Forum
Register Search Today's Posts Mark Forums Read

Real-Time PCR and Quantitative PCR Forum Real-Time PCR and Quantitative PCR Forum


Taqman based PCR

Taqman based PCR - Real-Time PCR and Quantitative PCR Forum

Taqman based PCR - Real-Time PCR and Quantitative PCR Forum


Reply
 
LinkBack Thread Tools Display Modes
  #1  
Old 03-17-2010, 10:57 AM
Pipette Filler
Points: 44, Level: 1 Points: 44, Level: 1 Points: 44, Level: 1
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Mar 2010
Posts: 1
Thanks: 0
Thanked 0 Times in 0 Posts
Default Taqman based PCR



Hello,

I am trying to amplify a PCR product of 120bp with a probe which has FAM at the 5' and BHQ1 at the 3' end. I am using the ABI Taqman Universal Master mix No amp No UNG for the amplification.

I use positive controls of cDNA for the real time. The temp used are 95 for 10 mins, 95 for 1 min, 48 for 40sec, and 60 for 1 min. 40 cycles of this. Data collection is done during the 60C cycle.

I have not been getting good results. The multicomponent data shows that there is more ROX signal than FAM signal and most of the samples positive by block based PCR and not positive during real time PCR.

At the end of the real time all the samples are run on a 1.5% agarose gel and strong bands are seen of the expected size. I am baffled as to what is going wrong.

The only thing I can suspect is the probe that I ordered, it could have degraded or not produced properly. How do I go about confirming that the probe is stable and has not been degraded.

thekid
Reply With Quote
Reply

Tags
based , pcr , taqman


Thread Tools
Display Modes

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
Human Cytome Project - Update 24 Jan. 2005 Peter Van Osta Cell Biology and Cell Culture 1 08-01-2010 02:18 PM
GNU units and units.dat; Units of Measurement and Unit Conversion James Redford Physics Forum 0 07-31-2005 12:08 PM
Human Cytome Project - Update 6 Jan. 2005 Peter Van Osta Cell Biology and Cell Culture 0 01-06-2005 11:18 AM
web/intranet based chemical inventory program Chemoventory 2.0 chemoventory Forum Chimica 0 03-01-2004 08:04 PM
Incredible find for Life based on Carbon (atomic number 6) Wild Stallion Physics Forum 2 07-09-2003 05:06 AM


All times are GMT. The time now is 04:59 PM.


Powered by vBulletin® Version 3.8.4
Copyright ©2000 - 2014, Jelsoft Enterprises Ltd.
Copyright 2005 - 2012 Molecular Station | All Rights Reserved
Page generated in 0.16620 seconds with 16 queries