Annual Report 2006

analysis using microsatellite markers showed
that a population of 40-60 adult females per
generation would be sufficient to conserve the
genetic diversity of the wild populations.
Minute pirate bug, is one of
the most effective predators of small insect
pests, such as thrips. We evaluated genetic
differentiations among wild populations of this
species using microsatellite DNA markers and
found that no difference among populations 100
km apart, suggesting that this species has large
gene pools in the field. We also analyzed
polymorphic DNA regions of mitochondrial and
nuclear DNAs amplified from Japanese
species. Phylogenetic analyses based on
nucleotide sequences showed that these DNA
regions can be used to examine genetic
interrelationships among species and strains.
Based on the results, we developed PCR
primers to amplify polymorphic DNAs from
many other insects.
Symbiotes
About 70,000 expressed sequence tags
were generated from the brown planthopper,
and new oligo-microarray
was designed based on a sequence clustering
analysis of the planthopper ESTs. A microarray
-analysis-room was newly set up for silkwormand
planthopper-arrays. The silkworm-array is
used by more than 12 domestic research
groups and planthopper-array by three groups.
RNAi protocol was successfully applied to the
planthoppers and is useful for the functional
analyses of the planthopper genes. Nucleotide
sequence analyses of ribosomal RNA and
mitochondria were performed in culicoides
species in Japan, and the species were identified
based on the nucleotide sequences.
We found two key amino-acid positions
affecting adaptability for heterologous overexpression
of termite cellulases (432 aminoacids,
a member of the glycoside-hydrolase
family 9) which generally resist against
recombinant production. By introducing aminoacid
mutations at the two positions, the termite
cellulases were over-expressed in .
The internal ribosome entry site (IRES) in
the intergenic region of intestine
virus binds with 80S ribosomes in the absence
of eukaryotic initiation factors. This indicates
that the IRES-mediated protein synthesis
would be possible in a complete reconstituted
system, containing 80S ribosome, mRNA,
tRNAs, aminoacyl tRNA synthetases (AARSs),
elongation factors, ATP, and GTP. We prepared
translation apparatus from eggs of brine
shrimp and reconstituted translation was
examined. Our AARSs were inactivated during
the purification procedure. If stable AARSs are
purified, eukaryotic reconstituted translation
system would be available with the IRES.
We found the infectivity against rice
yellow drawf-phytoplasma was different between
regional strains of green rice leafhopper,
and thereby were able to
build the experiment system of virulence
mechanism of phytoplasma insect pathogens.
It was examined by measuring the initial
quantity of entomopoxvirus
(AcEPV) fusolin gene in the ectoperitrophic
area of larvae with a method of realtime
quantitative PCR of the gene whether or
not a greater number of the AcEPV virions had
passed the peritrophic membrane (PM) after
the feeding of spindles mixed with spheroids
compared with that after the feeding of
spheroids only. The experimental results
indicated that a greater number of AcEPV
virions had passed through the disintegrated
sites generated in the PM by the spindles, and
that they had reached the ectoperitrophic area
and then had entered the midgut epithelium
within 6 hr after the administration of the
spindles.
Resistance of the silkworm to Cry1Ac
toxin of was related to
recessive major gene. We selected two
silkworm lines on the susceptibility against
entomogenous fungus,
infection. One line was resistant and the other
was susceptible. First filial generation (F1) of
two lines showed resistance and half of
backcrosses between F1 and susceptible line
showed resistance. These results suggested
that the resistance of the silkworm against
infection was related to dominant
major gene.