Marfan Syndrome mutation analysis ?

[Peter Van Osta]

Hello,


I am starting with the mutation analysis of our pool of Marfan patients, is 
there anybody reading this who is also working on mutation sreening of this 
disease ?

I am using exon-flanking intron primers for PCR amplification of the (65) 
exons of the FBN gene on genomic DNA, so I am not working on cDNA.

For the mutation-screening I intend to use Hydrolink MDE-gels for 
heteroduplex analysis, is there anyone who has used these gels for this gene ? 
What are the best conditions using this gel type (or another) to improve the 
detection rate in the fibrillin-gene ?
Is it necessary to use cooling for the gels, or is room temperature 
and low-power conditions sufficient for not disturbing the heterduplexes?

What is a good way for optimizing the formation of heteroduplexes ?
 

Thanks in advance for your help,




Peter VAN OSTA, MD

Centrum Medische Genetica
Universitair Ziekenhuis Gent (UZG) - 0 K 5
De Pintelaan 185
B-9000 Gent
BELGIUM

email: pvanosta at allserv.rug.ac.be
tel. +32-9-240 36 03
fax. +32-9-240 49 70