Page 203 - PC2019 Program & Proceedings

PLANT CANADA 2019

S191. An antisense oligoRIBO-11 fragment (contact DNA insecticide) penetrates through the
integuments into the cells of gypsy moth larvae (Lymantria dispar L.)
1
1*
1
3
Oberemok, V. ; K. Laikova ; I. Novikov ; N. Galchinsky ; R. Useinov , Y. Plugatar
1
2
1 V.I. Vernadsky Crimean Federal University
2 Research Institute of Agriculture of Crimea
3 Nikita Botanical Gardens—National Scientific Centre RAS
We developed the oligoRIBO-11 fragment (5'-TGCGTTCGAAA-3') (0.9 nmol/larva) and used it as an
antisense DNA oligonucleotide (DNA insecticide) to penetrate through the integuments into the cells of
gypsy moth larvae, a major insect pest of hardwood trees. Studies were then conducted using a variety of
mass spectrometric method – matrix-activated laser desorption/ionization (MALDI).

After the oligoRIBO-11 fragment was applied to the larvae, MALDI registered the penetration of the
fragment into the insect cells at 30 minutes post-treatment (peak at 3360.1 Da) and detected a significant
response to the applied oligonucleotide 60 minutes post-treatment. In the control group, a peak
characteristic of the oligoRIBO-11 fragment was not found. In addition, in the control group, the profile
of the recorded peaks (peaks at 2558.2 Da, 2582.2 Da, and 2910.5 Da) differed noticeably from those of
the experimental groups. This indicates that the oligonucleotide not only entered the insect cells, but also
that the synthesis of new substances (many of them were oligonucleotides) in response to the applied
DNA fragment occurred. Many new peaks were obtained in the diagram for samples 60 minutes post-
treatment with the oligoRIBO-11 fragment (peaks at 3604 Da, 4267.2 Da, 5509 Da, 7151 Da, 7737.1 Da,
8568.5 Da, and 9389.9 Da).

This provides compelling evidence that DNA insecticides are able to penetrate the integuments of the
gypsy moth larvae, triggering an active cell response.

Volodymyr Oberemok (genepcr@mail.ru)

S192. Transcriptomic analysis of red-berried grapevine infected with Grapevine leafroll-associated
virus 3
*
Song, Y. ; R. Hanner; B. Meng
University of Guelph

Grapevine leafroll-associated virus-3 (GLRaV-3) is the major virus associated with grapevine leafroll
complex and has a worldwide prevalence. The impact of GLRaV-3 infection on vine health, yield and
fruit quality at the molecular level has yet to be characterized. It is believed that GLRaV-3 infection is
associated with expressional changes of select genes in grapevine that are involved in biological pathways
fundamental for grapevine growth, development, and fruity quality. Red-berried grapevines (Vitis vinifera
cv. Cabernet franc) infected and uninfected with GLRaV-3 were identified using virus-specific primer
with reverse transcription polymerase chain reaction (RT-PCR). Transcriptome analysis by RNA
sequencing was carried out on leaves of identified grapevines at the harvest stage. Differentially
expressed genes (DEGs) suggested repressed photosynthetic activity with enhanced sink activity in
infected leaves, indicating source-to-sink transition in response to viral infection. Activation of defence
mechanisms was suggested by DEGs including increased biosynthesis of flavonoids, lignin, amino acids,
and differentially regulated biosynthesis and signalling of hormones. Our results from transcriptomic
analysis on red-berried grapevine leaf at harvest suggest down-regulation of biological processes that are
fundamental to plant growth and development while up-regulation of genes related to defense
mechanisms in responses to GLRaV-3 infection. Our ultimate goal is to understand the impact of
GLRaV-3 infection on global gene expression through transcriptome analysis of leaves and berries
collected at different growth stages.

Yashu Song (yashu@uoguelph.ca)

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