Go Back   Science Forums Biology Forum Molecular Biology Forum Physics Chemistry Forum > Molecular Research Topics Forum > Protocols and Methods Forum
Register Search Today's Posts Mark Forums Read

Protocols and Methods Forum Post Any Protocol, Method, Technique, Procedure or Tips / Troubleshooting for any Molecular Biology Technique.


AFLP low yield

AFLP low yield - Protocols and Methods Forum

AFLP low yield - Post Any Protocol, Method, Technique, Procedure or Tips / Troubleshooting for any Molecular Biology Technique.


Reply
 
LinkBack Thread Tools Display Modes
  #1  
Old 09-06-2006, 05:38 PM
aria johnson
Guest
 
Posts: n/a
Default AFLP low yield



Hi,

I am familiar with microsatellites, but currently trying to assess hylid DNA
using AFLP's and need advice on how to increase the yield of my 2nd amp
product, while improving the discrete bands that are barely visible in the 2
nd amp. smear.

When I run the product of the 2nd amplification ( on a 6% polyNAT gel) I get
a long (100bp - 800bp), faint smear with bands visible in the smear. Both
the lack of intensity of the product and the fact that I am not getting a
clean discrete banding pattern is unexpected because a) this is the 2nd amp.
with primers designed to increase specificity b) my positive control
(lambda) has a clear, bright, banding pattern (no smearing) . Could it be
that my annealing T is too low (not stringent enough), thereby producing
many fragments of varying sizes in addition to the ones I want? How can I
increase the overall product yield (more visible bands) while encouraging
selectivity in fragment amplification?



Loosing battle with AFLP
Reply With Quote
  #2  
Old 06-03-2009, 10:54 AM
Pipette Filler
Points: 52, Level: 1 Points: 52, Level: 1 Points: 52, Level: 1
Activity: 0% Activity: 0% Activity: 0%
 
Join Date: Jun 2009
Location: Hydreabad
Posts: 1
Thanks: 0
Thanked 0 Times in 0 Posts
Default Re: AFLP low yield

I had a similar issue, you can try increasing template quantity by increasing the number of cycles in pre select amplification, your product yield might increases

Last edited by ibtsaranya; 06-03-2009 at 11:34 AM.
Reply With Quote
Reply

Tags
aflp , low , yield


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
Poor yield: DNA purification by phenol/chloroform extraction + ethanol precipitation? amesy Protocols and Methods Forum 1 03-27-2009 11:18 PM
Poor Plasmid Prep Yield zek Molecular Cloning Forum 0 02-10-2009 07:45 PM
RNA yield per fly? Chris Jones Drosophila Forum 1 01-19-2006 08:32 AM
how to increase cosmid yield BoonChoon Protocols and Methods Forum 1 12-08-2004 05:12 PM
FW: AFLP Deanne Bell Protocols and Methods Forum 0 05-11-2004 06:23 PM


All times are GMT. The time now is 12:24 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 0.12213 seconds with 16 queries