Transgenesis in C. elegans - Caenorhabditis Elegans: Molecular Genetics and Development - page 169

Table III (Continued)

Technique Applications Considerations References

Cell-specific delivery of heat
shock

Sensitive spatial and temporal
control of gene expression
using a focused laser
microbeam

Requires laser apparatus and
ability to identify cells. Care
is required to avoid
damaging the induced cell(s)

Stringham et al.
(1992)

Temperature-sensitive mec-8-
dependent splicing

Permits controlled, temperature-
sensitive regulation of gene
expression, including RNAi-
sensitivity

Perdurance of mec-8 activity can
make precise regulation
difficult.
Splicing event requires low-
doses of MEC-8. Need to
work in mec-8 background

Calixto et al. (2010)

Selective depolarization of cells
by light stimulation
(''optogenetics'')

Activation of transgene-driven
light-sensitive proteins such
as channelrhodopsin-2
(ChR2) ( Nagel et al., 2003 ).

Light activation can be delivered
broadly, as only cells
expressing ChR2 will
become depolarized. Light-
sensitive channels that
respond to different
wavelengths can be used
simultaneously

Stirman et al. (2011)

Tissue-specific RNAi sensitivity Permits specific loss of gene

activity in a subset of cells

RNAi effectiveness can be
variable

Qadota et al. (2007)

identify which cells/tissues have inherited the array, cells carrying the wild-type
copy of a gene could be identified by tagging the gene with GFP, by including a
ubiquitous reporter such as sur-5::GFP ( Yochem and Herman, 2005 ), by including
rescue of ncl-1, whose function can be scored cell-autonomously ( Hedgecock and
Herman, 1995 ), or by using a nuclear-localized GFP::LacI to mark LacO sequences
present in the array ( Gonzalez-Serricchio and Sternberg, 2006 ) (discussed below).
By referring back to the C. elegans lineage ( Sulston et al., 1983 ), the researcher can
conclude which cell(s) lost the array in a particular animal, and, if this loss includes
tissues of interest, conclusions can be made about cell-autonomous and cell nonau-
tonomous functions. Finally, arrays can be specifically lost in the maternal germ line,
so that progeny animals can be produced that lack both maternal and zygotic
contributions of the gene ( Hunter and Kenyon, 1996 ). While this approach has been
immensely powerful, using array loss to examine tissue-specific gene expression
does have limitations, which include the sometimes-complex lineage analysis
required to understand emerging phenotypes.
A second technique for examining tissue-specific expression relies on the use of
tissue-specific promoters linked to one ' s gene of interest ( Table III ). While this
technique has also significantly contributed to our understanding of tissue-specific
gene expression, this analysis can be restricted by the limited availability of well-
characterized promoters.
A number of strategies for temporally and spatially controlling gene expression
have been recently developed ( Table IV ). In general, these techniques depend on