Deciphering Drosophila DNA

P(acman) -- a new method of introducing DNA into the genome of fruit
flies or Drosophila -- promises to transform the ability of scientists
to study the structure and function of virtually all the fly's genes,
and the method may be applicable to other frequently studied organisms
such as mice, said its Baylor College of Medicine developers in an
article in the current issue of the journal Science.

"P(acman) overcomes a key limitation of currently available methods
because it allows you to study large chunks of DNA in vivo," said Dr.
Hugo Bellen, professor of molecular and human genetics at Baylor College
of Medicine and director of the program in developmental biology. He is
also a Howard Hughes Medical Institute investigator. The new technique
allows researchers to study large genes and even gene complexes in the
fruit fly, which was not possible before.

P/phiC31 artificial chromosome for manipulation, or P(acman), combines
three recently developed technologies: a specially designed bacterial
artificial chromosome (BAC) that allows maintenance of large pieces of
DNA in bacteria, recombineering that allows the manipulation of large
pieces of DNA that can then be inserted into the genome of the fly at a
specific site using phiC31-mediated transgenesis.

It is a new technique with far-reaching promise, said Bellen.

P(acman) overcomes certain obstacles that have hampered research. It
allows the cloning of large pieces of DNA to be used to transform the
genome, and it permits that DNA to be inserted into specific places in
the genome. Bellen credits the report's first author, Koen J.T. Venken,
a graduate student in the BCM Program in Developmental Biology, with
putting the technologies together to come up with a new methodology in
the field.

Current technology has certain problems for researchers seeking to
understand the structure and function of genes, said Bellen. Often, when
scientists breed flies that lack a particular gene and then try to put
that gene back into the fly, it inserts itself randomly into the genetic
blueprint.

In some cases, it makes too much protein, and in others, too little. In
other instances, it may disrupt the message from another gene.

"You are really comparing apples and oranges when you do this," said
Bellen. The technique is also limited to small DNA chunks.

"Koen set out to develop a new transgenesis system using the three
techniques," said Bellen.

The bacterial artificial chromosome, or BAC, he used allows the
scientist to maintain large chunks of DNA in the bacteria, but it is
present in only one or few copies. However, the bacteria can be induced
to produce many copies of the DNA when needed.

Koen then integrated a technique called "recombineering" into the
strategy, which facilitates the scientist to clone large chunks of DNA
and subsequently allows them to make specific mutations anywhere he or
she wants in the gene.

The third technique allows the researcher to pinpoint where he or she
wants to the mutant gene to go in the genetic blueprint of the fly,
eliminating the apples-and-oranges problem. This third technique --
phiC31 -- works also in mouse and human cells, implying that this new
technique could be used in those cells as well.

Others who contributed to this report include Yuchun He, also of BCM,
and Dr. Roger A. Hoskins of Lawrence Berkeley National Laboratory in
Berkeley, California.

The work was supported by the National Institutes of Health and the
Howard Hughes Medical Institute.

Original Article
<http://www.sciencedaily.com/releases/2006/11/061130191541.htm>