VUV MASS SPECTROMETER SHOWS PROMISE                 
   
   One of the more unlikely players in the DNA sequencing game is Winston
   Chen of ORNL's Health and Safety Research Division. "We had been using
   lasers to do research in the physical sciences," Chen explains, "and we
   wanted to see if we could expand into biological and environmental
   research."          
   
   Chen, Marvin G. Payne, also of the Health and Safety Research Division,
   and Bruce Jacobson of the Biology Division are developing a new
   high-resolution mass spectrometer equipped with a vacuum ultraviolet
   (VUV) photoionizer to sequence ferrocene-tagged DNA. This approach could
   eliminate the need for both gel electrophoresis and radioactive tagging
   while sequencing DNA segments that are thousands of bases long.            
         
   
   In this procedure, a primer (a small molecule to which DNA bases are
   added) is labeled with an organometallic compound, such as ferrocene.
   Then new DNA segments that terminate at each occurrence of a particular
   DNA base (A, G, C, or T) are built up on the primer using the original
   DNA as a template.        
   
   To ionize these fragile molecules without breaking them up, the group is
   using a technique known as laser desorption from a cryogenic matrix. In
   this technique, DNA segments are placed on a thin layer of solid argon
   that has been cooled to about 20 K on a cryogenic copper finger. A laser
   is then used to warm the finger to 100 K, causing both the argon and the
   DNA to vaporize and flow into the ionizer. Argon's low vaporization
   point makes fragmentation of the DNA segments during ionization very
   unlikely. Chen and his colleagues have sequenced DNA segments up to 34
   bases long with a single laser pulse using this method. The system they
   are developing has 50 times the resolution of the current system and is
   expected to be able to sequence DNA segments up to 1000 to 100,000 bases
   long.        
   
   When the DNA segments reach the ionizer in the new system, they will be
   ionized by a weak vacuum ultraviolet (VUV) beam. Preliminary studies on
   other fragile organic molecules indicate this process is very unlikely
   to result in fragmentation of the DNA segments. Also, this method has
   the advantage of being energetic enough to ionize ferrocene-tagged DNA,
   but not energetic enough to ionize untagged DNA or other possible
   impurities.         
   
   Finally, DNA ions resulting from VUV ionization are sent on to a
   high-resolution time-of-flight mass spectrometer where their masses and
   sequences are determined. "This type of mass spectrometer makes possible
   a greater separation between pieces of DNA at the detector," Chen says,
   "and makes it easier to distinguish between segments that differ by only
   one base."          
   
   The value of this novel technique is its potential for improving
   efficiency on several fronts. "This method has the potential for
   sequencing 10,000 times as many bases as gel electrophoresis," says
   Chen. "Our technique could also significantly reduce preparation time
   and processing time and avoids the use of gels, radioactive materials,
   and fluorescent dyes.            
   
   "For instance, with gel methods the speed of molecules can be measured
   in meters per hour. Using the vacuum method, the speed can be measured
   in kilometers per second. As a result, we may be able to do separations
   that used to take several hours using gel electrophoresis in less than
   one second.
   

   ------------------------------------------------------------------------
   
   Please send us your comments.
   
   Date Posted:  1/10/94  (ktb)