Transgenesis in C. elegans - Caenorhabditis Elegans: Molecular Genetics and Development - page 178

D. Transgenics in Other Nematode Species
With the recent availability of genome sequences for other nematodes, researchers
may wish to perform gene manipulations in these species. The closely related
C. briggsae is the most frequent choice for comparative work, perhaps because it
is hermaphroditic like C. elegans (most others are male-female), and because its
genome sequence is of very high quality ( Stein et al., 2003 ). Like C. elegans,
C. briggsae can be made transgenic by microinjection with the use of rol-6D or
rescue of mutants. Mutants in unc-119 have been made available that permit the use
of microparticle bombardment ( Zhao et al., 2010 ). There are Mos insertions that
have been made in C. briggsae but the MosSCI approach is still being developed
(Marie Delattre, personal communication). In the more distant nematode Pristionchus
pacificus, also a hermaphroditic species, transgenics can be made, though with some
difficulty ( Schlager et al., 2009 ), using an adaptation of the protocol for using
complex arrays in C. elegans ( Kelly et al., 1997 ). Routine transgenesis in male-
female nematode species has not been developed, although in principle coinjection of
a dominant marker that does not affect male mating, such as a GFP reporter, could be
used to mark transgenics. The rol-6D phenotype compromises male mating, which
would make maintenance of homozygous transgenic integrants more difficult. As
well, the basis for identification of transgenics in bombardment and MosSCI - unc-
119 rescue - would be impossible in male-female species because unc-119 blocks
male mating. Microparticle bombardment may be the best possibility for transgenesis
in other species, if a system can be devised to identify rare transgenics. One promising
breakthrough, a transformation strategy that depends on conferring drug resistance,
will likely make it simpler to generate transgenics in a large number of nematode
species ( Giordano-Santini et al., 2010; Semple et al.,2010 ).

V. Perspectives: What Lies on the Horizon?

While recent research has added to an already rich suite of applications for
transgenesis in C. elegans, there are some technologies, used in other systems, which
are still being developed or refined in worms. One essential technique for studying
gene function is a simple, reproducible method for knocking out, tagging, or other-
wise manipulating a gene at its endogenous locus. Several recently developed
methods for creating lines carrying homologous integrations, using negative/posi-
tive selection after microparticle bombardment or MosSCI, promise to make homol-
ogous, targeted modifications the standard in C. elegans ( Frokjaer-Jensen et al.,

2008; Vazquez-Manrique et al., 2010 ).
A second essential technique that would permit more sophisticated analysis of
gene function is one that promotes or prevents gene expression in a precise spatial or
temporal pattern, similar to the Drosophila GAL4 system. In principle, several
recently developed techniques described in the mosaic analysis section of this
chapter could achieve this aim ( Table III ). Particularly promising are those techni-
ques that create strains that confer specific expression or inhibition of any