dna array

[Seng-Lai Tan]

Hi,

we have been using the high density array hybridization (HDAH) technique
to study differential gene expression.  we used an automated bacterial
colony picking and arraying tool to array bacterial clones onto nylon
membranes.  The membranes were places on LB agar and incubated until
colonies were uniform in size.  The colonies were lysed, denatured, and
neutralized directly on the filters, which were then hybridized with
radio-labeled complex cDNA probes.  

everything was working fine; we could see nice differential expression on
our filters when hybridized with complex cDNA probes generated from mRNAs
of different biological samples.  However, lately we have not been able to
reproducible our results, i.e. we can no longer see any differential
expression on our filters even for genes that we know should show big
differences.  we do not know what's wrong as we did not change the
procedures for these experiments.  we wonder if anyone out there has
encountered similar experience.  Any advice and/or suggestion that you
may have is very much appreciated.

Sincerely,                                  
Thomas S-L. Tan                   
Department of Microbiology	  
Box 357242			  
Health Sciences Center            
University of Washington          
School of Medicine	           
Seattle, WA 98195 		  
                                  
Phone:  206-543-5915			  
FAX: 	206-543-8297
E-mail: sltan at dante.u.washington.edu