Emerging Technologies

Articles:

• Padlock Probes Detect Single Base Changes
• AFM Imaging of DNA-Bound Proteins
• Mutation Databases
• DOE Human Genome Program Contractor-Grantee Workshop V (1996 abstracts)
• DOE Human Genome Program Contractor-Grantee Workshop VI (1997 abstracts)

(1) Paul M. Lizardi and David C. Ward. "FISH with a twist." Nature Genetics 16: 217-218, 1997.
(2) Mats Nilsson et al. "Padlock probes reveal single-nucleotide differences, parent of origin and in situ distribution of centromeric sequences in human chromosomes 13 and 21. Nature Genetics 16: 252-255, 1997.

Atomic Force Microscopy (AFM) Imaging of DNA-Bound Proteins

Oak Ridge National Laboratory (ORNL) scientists David Allison and Mitchel Doktycz are developing a new imaging approach for DNA mapping that may also aid in the rapid identification mutations. Using the atomic force microscope they have been able to identify EcoRI endonuclease site-specifically bound to the appropriate sites on individual DNA molecules. This was accomplished using a mutant enzyme that binds to the specific nucleotide sequence GAATTC but, unlike the wildtype EcoRI endonuclease, does not cleave the DNA. The resolution of the technique was determined by imaging well characterized plasmids with one or two EcoRI sites. They found excellent agreement between the predicted and measured values, with a deviation of about 1% when measuring 10 molecules or fragments, which is better than can be obtained with gel electrophoretic analysis. Recently they have mapped lambda bacteriophage DNA, a well characterized larger molecule of ~50,000 base pairs with five EcoRI sites, and cosmid clones of mouse genomic DNA with similar results.

Future applications of this new imaging technology will be the localization of other important site-specific DNA-protein interactions. Often this information is inaccessible or cumbersome to obtain using other methods of analysis. For example, transcription factors that recognize specific sequences in gene promoter regions could be imaged. This would allow the identification of gene start regions, thereby identifying genes, on cloned DNA molecules. A possible application of AFM imaging to Mutation Research that immediately comes to mind is its potential for identifying the number and locations of mutations in genomic DNA by imaging repair enzymes bound to altered DNA base sequences.

Mutation Databases

For some years, researchers have created and maintained databases for their own research purposes and have often shared them in review articles. More recently, these databases have been placed on the WWW allowing easy distribution of the database. Necessarily those databases have been created in isolation and thus, content is not uniform and comparisons are not easy. An initiative has begun to obviate this problem by working towards up-to-date lists of mutations available on the WWW. This initiative, supported by HUGO, has already had two International meetings which has set up working parties and is having its third meeting on the 29th October, 1996. It is expected that an association "The Mutation Database Association" will be formed of interested people at that time. These databases are not intended to be human only, but this is where the initiative has begun.

More details can be located on the Mutation Database Web site

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Monday, November 17, 2003

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