Nematodes of the genus Meloidogyne are known as Âroot-knot nematodes due
to the characteristic knots or galls found on the roots of infected plants. Root-knot
nematodes attack over 2000 species of plants and cause over 80 billion dollars lost
annually. Giant cells are feeding sites of root-knot nematodes and are enlarged
multinuclear cells induced by the nematodes in susceptible host roots and that function
to provide nutrients to the nematode. This thesis presents data on two techniques of
studying the nuclear condition in giant cells. Colchicine was used to arrest mitosis in
giant cells in a previous study. Here we test the effect of colchicine on nematode
activity. The results showed that colchicine did not affect nematode egg hatch, juvenile
activity, or hatch of eggs produced by treated juveniles. These results confirm that
colchicine can be used to arrest mitosis in giant cells without affecting the nematode
parasite.
A major obstacle to the study of giant cells is collecting tissue samples that are
specific to giant cells. Laser capture microdissection (LCM) is a technique that allows
one to sample a single giant cell. A focused laser beam was used to collect samples of
giant cell cytoplasm from fixed and sectioned tissues. RNA was then extracted from
those isolated samples. Using three tomato genes as test samples, specific primers were
designed to measure expression level of Rb7, LHA4, and HXK1 gene by Real-Time PCR.
Expression of LHA4 and Rb7 increased with time after inoculation, and immature giant
cells reached levels that were 3 and 6 times, respectively, that of cortical cells, but which
were not different from root meristem cells. Expression of HXK1 did not change with
time after inoculation and has the same level of that in root tip and cortical tissues.
These data confirmed that the techniques of LCM coupled with RT-real-time PCR can
be used to quantitate expression of genes at different stages of giant cell development
without contamination from surrounding cells.
| Identifer | oai:union.ndltd.org:tamu.edu/oai:repository.tamu.edu:1969.1/3243 |
| Date | 12 April 2006 |
| Creators | He, Bin |
| Contributors | Starr, James L. |
| Publisher | Texas A&M University |
| Source Sets | Texas A and M University |
| Language | en_US |
| Detected Language | English |
| Type | Book, Thesis, Electronic Thesis, text |
| Format | 232083 bytes, electronic, application/pdf, born digital |
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