Hi all,

I am attempting to extract DNA for sequencing from formalin fixed
and stored larval fish tissues.  I was curious if anyone had any good
protocols or tips and tricks to help me accomplish this task.  My ultimate
goal is to get a long enough sequence to be able to compare to a known
adult sequence and confirm identification.

Thanks so much,

Carie Bikson
UW Fish Collection
University of Washington, Seattle, WA
cbikson@uw.edu
 

Responses:

 

·        
Palero, Hall, Clark, Johnston, Mackenzie-Dodds
and Thatje. 2010. DNA extraction from formalin-fixed tissue: new light from the
deep sea. Scientia Marina
74(3): 465-470.

 

·        
Fang, Wan and Fujihara. 2002. Formalin removal
from archival tissue by critical point drying. Biotechniques 33(3): 604-611.

 

·        
Shedlock, Haygood, Pietsch and Bentzen. 1997.
Enhanced DNA extraction and PCR amplification of mitochondrial genes from
formalin-fixed museum specimens. Biotechniques 22: 394-396

 

·        
Schander and Halanych. 2003. DNA, PCR and
formalinized animal tissue – a short review and protocols. Org. Divers. Evol.
3: 195-205.

 

·        
Austin and Melville. 2006. Incorporating historical
museum specimens into molecular systematic and conservation genetics research.
Molecular Ecology Notes 6: 1089-1092.

 

·        
Path to Effective Recovering of DNA from
Formalin-Fixed Biological Samples in Natural History Collections: Workshop
Summary (2006). Board on Life Sciences

http://www.nap.edu/catalog.php?record_id=11712#toc

 

·        
Chase, Etter, Rex and Quattro. 1998. Extraction
and amplification of mitochondrial DNA from formalin-fixed deep-sea mollusks.
Biotechniques 24(2): 243-247.

 

·        
QIAamp DNA FFPE Tissue Kit. Qiagen

http://www.qiagen.com/Products/QIAampDNAFFPETissueKit.aspx

 

·        
Depends on how long they were in formalin. 
I had success by using a set of successive baths for tissue pieces.  I
started with a day in TE, then moved to 50% EtOH/50% TE and then each day
increased the EtOH 10% until the tissues were in 95% EtOH.  I got
amplifiable DNA out of some Brazilian holotypes that way.

Peter Wimberger

 

·        
NucleoSpin Tissue XS Kits. ClonTech

http://www.clontech.com/products/detail.asp?product_id=10537&tabno=2

 

·        
GenomePlex Whole Genome Amplification Kit.
Sigma-Aldrich.

http://www.sigmaaldrich.com/life-science/molecular-biology/automation/whole-genome-amplification.html

 

·        
Gilbert, Haselkom, Bunce, Sanchez, Lucas et al.
2007. The Isolation of Nucleic Acids from Fixed, Paraffin-Embedded Tissues –
Which Methods Are Useful When?. PLoS ONE 2(6): e537.
doi:10.1371/journal.pone.0000537

 

·        
Robertson, LeDuc, LeDuc and Morin. 2007.
Extraction of DNA From Formalin-Fixed Cetacean Tissues. NOAA Technical Memorandum
NMFS. February 2007. NOAA-TM-NMFS-SWFSC-400.

Carie Bikson
University of Washington
School of Aquatic and Fishery Sciences
Box 355020
Seattle, WA 98195
Fisheries Teaching and Research Room 005
(913)205-7383
carie_berry@yahoo.com
cbikson@u.washington.edu

Carie Bikson <carie_berry@yahoo.com>