Agriculture Reference
In-Depth Information
gels. These bands can be visually scored and compared for the differences in inten-
sity for detecting transcripts expressed differentially
in plants
and for comparison
these will be eluted and reamplified with appropriate cDNA-AFLP primers. This is
followed by sequencing and then the products are screened using public databases
for finding homologous sequences with significant alignment. Using the BLAST
analysis, sequences identical to crop transcripts, transcripts homologous to known
plant sequences in UNIPORT KB Swiss-Prot or TrEMBL http://www.expasy.ch/
sprot/databases or NCBI databases (http://blast.ncbi.nlm.nih.gov/Blast.cgi) and
transcripts homologous to known
Fusarium
sp. sequences considered as express-
ing
in planta
shall be identified. The transcripts, which have no matches in any
of the databases, shall also be seen; these may represent transcripts that currently
lack functional annotations. Further the expression pattern and clustering of crop
transcripts shall be done to overview differences between infected and control. The
functional annotation of each transcript can be done with help of Gene Ontology
Database http://www.geneontology.org and through carefully analyzing the scien-
tific literature. Further for the identification of
Fusarium
genes expressed in crop
plant during infection we can include sequences from all
Fusarium
sp available in
public databases. Finally, the fungal transcripts differentially expressed among the
strains shall be identified when grown
in vitro
searching the
Fusarium
database (Fu-
sarium Comparative Database 2). These studies will provide with information re-
garding transcriptional changes in crops upon
Fusarium
pathogenesis and also if the
wilting symptoms are derived from active plant response besides the infection. The
studies shall generate information regarding
In planta
expressed pathogen-derived
transcripts during the infection processes which are related potentially to virulence
functions as well as the
in vitro
expressed transcripts (expressed differentially be-
tween strains). These will further provide the sequences, which will be helpful for
distinguishing between races. For the dissipation of information the sequence data
generated from these studies shall be deposited in Gene Bank (Gene Expression
Omnibus (GEO) database (http://www.ncbi.nlm.nih.gov/geo). The second impor-
tant concern is related to Identification of the target regions in the fungal genome
for probe generation for their use in phylogeny of
Fusarium
sp. Further, the use of
a comprehensive EST collection in
Fusarium
for SSR identification shall help to
develop EST-SSR markers for genetic mapping which will be extremely useful for
diagnostics and research concerned with fungal biology, ecology, and genetics. The
analysis will yield the frequency, type and distribution of SSR motifs in ESTs de-
rived from
Fusarium
sp. The Perl script MIcroSAtelitte (MISA) (Thiel et al.
2003
)
can be used to identify SSRs in the
Fusarium spp.
EST sequences. These data can
be used to perform comparative analysis of SSR motif polymorphisms between
allelic sequences, and orthologous sequences to conduct and finally to identify
functionally associated EST-SSR markers for application in comparative genomics.
There is a need to study and focus on the development of EST-SSRs. Due to the lack
of sufficient markers for
Fusarium
sp, it becomes necessary to develop enough mo-
lecular markers for potential use in
Fusarium
sp. pathogenicity. With the develop-
ment of
Fusarium
database/projects a vast amount of available EST sequence data
has been generated. The screening for repeat motifs (perfect and imperfect) can be
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