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TRiP
>> Approach - Overview
Targeted transgenic RNA
The approach used by the TRiP is to generate transgenic
animals with an RNAi hairpin under UAS-Gal4 control. The
hairpin-containing transgenes are inserted via site-specific
recombination into genomic loci known to be optimal for
expression. Specific information follows.
Advantages of the TRiP Stocks and VALIUM Vectors
The targeted method to generate hairpin lines has many
practical advantages over P-element based methods: 1. the
frequency of recovering transformants using the integrase
method, either following co-injection of integrase mRNA or by
injecting into the nanos-integrase strain, is almost five-fold
higher than in conventional P-element transformation; 2.
establishment of the lines is greatly facilitated as no mapping
of the transformants to a specific chromosome is needed; and 3.
unlike P-element-based methods, insertions into the attP
landing site are homozygous viable.
Further, since the efficacy of the transgenic RNAi
technology depends upon the level of expression of the
UAS-driven constructs, the VALIUM constructs with their modular
number of UAS copies allow the generation of a phenotypic
series. From the original 10XUAS construct, a 5XUAS derivative
can be recovered, and because the attP-containing chromosome
can be homozygosed, it is also possible to generate both 15XUAS
and 20XUAS combinations (Ni et al., 2008). The ability to
generate a phenotypic series from 5XUAS to 20XUAS may prove
useful, in particular, when variation of the gene expression
dosage is important for phenotypic studies of pleiotropic
genes. To generate different levels of RNAi knockdown, the
features described above can be used together with other means
to vary expression (different Gal4 lines of different strength,
temperature) or processing (with coexpression of UAS-Dcr2,
Dietzl et al., 2007) of the hairpin construct.
Design and Production
Selected References
Ni, J-Q., Markstein, M., Binari, R., Pfeiffer,
B., Liu, L-P., Villalta, C., Booker, M., Perkins, L. A., and
Perrimon, N. (2008) Vector and Parameters for Targeted
Transgenic RNAi in Drosophila melanogaster. Nature Methods 5,
49-51.  
Dietzl, G. et al. (2007) A genome-wide
transgenic RNAi library for conditional gene inactivation in
Drosophila. Nature 448, 151-156.
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