Biology Reference
In-Depth Information
effects on gene expression. Genes introduced by TE and viral vectors insert more
or less randomly into the chromosomes, making it difficult to predict how well the
transgene will be expressed. The use of zinc-finger nucleases, homing endonucle-
ases, and TAL effector nucleases (TALENs) to genetically modify arthropod genomes
in a specific manner was described in Chapter 9. To date, these methods are being
used as tools to evaluate gene function (
Wright et al. 2006, Miller et al. 2007,
Beumer et al. 2008, Traver et al. 2009
,
Takasu et al. 2010
,
Urnov et al. 2010
,
Carroll
2011, Windbichler et al. 2011, Watanabe et al. 2012
), but could be used to develop
genetically modified arthropods for pest-management programs. At present, how-
ever, the many issues regarding their stability and their potential risks and relevant
regulations have not been resolved. However, the potential risks of HT by these
methods could be greatly reduced compared with the use of TE vectors.
Another method for accomplishing precise insertion is based on a system found
in the yeast
Saccharomyces cerevisiae
. A gene for yeast recombinase, FLP recombi-
nase, and two inverted recombination target sites (FRTs) that are specifically rec-
ognized by the FLP recombinase have been cloned. The FLP-FRT system has been
modified to insert DNA into a specific site in a
Drosophila
chromosome (
Konsolaki
et al. 1992, Simpson 1993, Golic et al. 1997
). If the FRT sites are inserted into other
insects, the system could reduce concerns about unstable transformation that are
elicited by TE vectors. Because a stable FRT site must be present in the genome, a
number of different lines carrying FRT sites in different chromosomal locations will
have to be evaluated to determine which site permits better expression of the intro-
duced genes. The FRT system is introduced into
D. melanogaster
using TE vectors, so
vector sequences may have to be removed to preclude subsequent movement.
14.5.6 No Vectors
A few experiments have delivered linear or circular plasmid DNA into the
genomes of arthropods without using a specific vector (
Presnail and Hoy 1992,
1996
). This approach has the advantage of eliminating potential risks of intro-
ducing TE sequences into the insect genome, which could result in increased sta-
bility of the inserted genes in the genome. It assumes that the inserted gene is
no more likely than any other gene to be moved by endogenous TEs or viruses.
How integration occurs is unknown, but could involve nonhomologous recombi-
nation or DNA-repair mechanisms.
14.5.7 RNAi to Control Pests
RNAi was described in Chapter 9 as a method for analyzing gene function in
D. melanogaster
. However, it has potential for use in managing pest arthro-
pods, as well (
De la Fuente et al. 2007, Whyard et al. 2009, Belles 2010, Huvenne
