An attempt has been
made to prioritize these items, but they are all considered to be high
of the remaining genomic sequence.
are best done with a complete genomic sequence.
b. A complete,
unigene set of full-length sequenced cDNAs.
The cDNAs will be
useful for confirming the expression of predicted genes and patterns
of alternative pre-mRNA splicing. They will also be used in microarrays
for monitoring genome-wide patterns of transcription, for systematic
RNA-mediated interference (RNAi) experiments aimed at transient inactivation
of specific gene expression, for two-hybrid libraries, and for in vitro
synthesis of protein products to be used in biochemical experiments.
Both cDNAs and primer pairs for each cDNA should be available for distribution.
Cost: $3-8 Million
Time: 3 years
c. Central resource
for generating gene knockouts (KOs).
We strongly endorse
the current pilot project for generating gene knockouts. Top priority
should be given to requests from academic researchers for specific KOs,
but ultimately KOs of every gene are desired. We also encourage efforts
to improve the technology, including the adoption of systematic procedures
for outcrossing and homozygosing (or balancing) each KO. A method for
targeted gene disruption by homologous recombination would be highly
on technology development
d. Make microarray
technology available to community.
We need a central
resource to generate whole genome chips and distribute them to the community
at an affordable price. The resource might also provide software to
analyze data and generate a publicly-accessible gene expression database.
Cost: $1.2 Million
Time: 3 years
for initial phase
e. Complete genomic
sequence of C. briggsae and possibly another nematode.
Comparisons of C.
elegans and C. briggsae, which diverged from each other at
least 20 million years ago, have proved to be good predictors of gene
structure and functionally important regions, especially regions that
regulate transcription. By identifying conserved transcriptional regulatory
regions, it may be possible to assemble regulatory cascades simply by
searching whole genome sequence for common (and conserved) transcription
factor binding sites. There may also be much to learn from sequencing
another, more distantly related, nematode, perhaps a parasite.
Cost: $25 Million
for C. briggsae
Time: 5 years?
f. Expanded stock
center and creation of a vector resource.
A significant increase
in genetic stocks will require increased resources for the C. elegans
stock center (the Caenorhabditis Genetics Center, or CGC). We also recommend
the creation of a center for distribution of commonly-used vectors.
Cost: $0.3 Million
Time: As long
g. High density
map of DNA dimorphisms for physically mapping genes defined by mutation.
Relating gene to
function will continue to involve analyzing mutant phenotypes, characterizing
genes, gene products and interactions among genes and gene products.
Genetic mapping strains with sequenced polymorphisms every few kb would
greatly aid continuing efforts to identify the DNA sequence of genes
defined solely by mutant phenotypes. Technology to facilitate displaying
large numbers of polymorphisms, possibly using microarrays, is also
Cost: $1 Million,
depending on technology development
Time: 3 years
h. Creation of
transgenic lines carrying GFP fusions to every gene.
A central resource
is needed to generate GFP fusions, create stable transgenic lines carrying
these fusions, and analyze the expression patterns as close to cell
resolution as possible. These patterns should be catalogued in a publicly-accessible
Cost: $10 Million
Time: 5 years
i. Methods and
We recommend support
for investigator-initiated research projects aimed at improving or creating
technologies of use to the community. These would include more efficient
transformation, more efficient gene KOs, methods for targeted gene replacement,
better regulation of transgenic gene expression, ability to do cell
culture, better electrophysiology, and better methods for systematic
phenotypic characterization, including automated methods for following
It should perhaps
be more widely advertised that NIH enthusiastically funds technology
development and that study sections can be very receptive to applications
that concentrate on technology development if the proposed technology
is shown to have a benefit, if the relevant expertise, both biological
and technical, is apparent (collaborations may be useful), and if the
proposal is well designed. Two other suggestions for enhancing technology
development through standard grants would be, first, to make clear to
study sections that significant progress in technology development may
not result in prominent publications, and second, to restore the five-year
Cost: R01 and