GENETIC TRANSFORMATION OF TOBACCO WITH Hs1(pro-1)
GENE AND EVALUATION OF NEMATODE RESISTANCE IN TRANSGENIC PLANTS
Ms., Department of Biotechnology
Supervisor: Prof. Dr. Huseyin Avni Oktem
Co-Supervisor: Prof. Dr. Halil Elekcioglu
September, 2001, 82 pages
Plant-parasitic nematodes constitute 10% of all
the nematodes and they are the major animal parasites of plants.
They have a very broad host range and they cause an estimated 10 to
25% annual reduction in crop yield along with a global economic
effect on crops of more than $100 billion annually.
This study aimed to genetically transform tobacco
plant with a nematode resistance gene Hs1pro-1 via Agrobacterium
mediated gene transfer technology, in order to develop transgenic
tobacco plants conferring resistance to nematode infection.
For this purpose, an Agrobacterium strain, EHA105
has been transformed with the plasmid, pDNEM33 bearing the Hs1pro-1
electroporation. The newly formed bacterial culture, EHA105::pDNEM33
was used in the plant transformation studies.
After the transformation studies, putative
transgenic plants were regenerated under the selection of
Phosphinothricin (PPT) with varying concentrations and several types
of analyses were carried out on the regenerated plants.
The leaf disc assay and colour viability test for
the plants under selection of PPT (5mg/L) revealed positive results
compared to the wild type control plants. The PCR analysis and the
Southern Blot Hybridization results also indicated that the gene,
Hs1pro-1 was stably inserted within the plant genome.
Finally, bioassays of the F1 seeds were performed
with two different nematode species, Meloidogyne incognita and M.
javanica. The roots of both the wild type and transgenic tobacco
plants were examined 20-days post infection and evaluated according
to 0-10 Gall Index Scale, the wild type and transgenic samples were
compared by one-way ANOVA test and found out to be statistically
different. The results indicated that, transgenic tobacco seedlings
were less affected by nematode infection compared to the wild type
Keywords: plant-parasitic nematodes, nematode
resistance, Hs1pro-1 gene, transgenic tobacco, Agrobacterium
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