ANNUAL REPORT - Department of Biotechnology

producing low lignin varieties. A full length clone of
OMT ( ~ 1.1 Kb) was isolated by screening of cDNA
library of Leucaena leucocephala and sequenced.
On blast analysis, the sequence exhibited homology
to reported OMT sequences in the data bank
indicating authenticity of our clone. Leucaena
leucpcephala was transformed with an antisense
OMT gene construct from aspen and analysis of the
transformant was done. The integration of a
heterologous antisense OMT gene construct in
transformed plants led to a maximum of 60%
reduction in OMT activity relative to control. The
evaluation of total lignin content by Klason method
revealed a maximum of 28% reduction.
In the area of horticulture, studies were focused on
development of protocols for generation of superior
planting material, molecular characterization of
germplasm and development of improved varieties
for increased shelf life and disease resistance. Under
the apple network programme, protocols for
micropropagation of important root stocks were
standardized and mass produced and nearly 10,000
root stocks have been planted with approximately 85
to 90% success. Protocol for root stock Merton 793 &
M26 has also been standardized and approximately
15,000 plants have been produced, which would be
field planted shortly. Germplasm assessment and
molecular characterization for apple was
successfully conducted during the year. 119 superior
cultivars were collected from 13 orchards and
morphological descriptors prepared for 80 cultivars
with molecular profiles generated for 78 and
cytological studies completed for 56 genotypes with a
significant achievements of identification of a scab
resistant cultivar. The microsatellite markers are
being developed for specific traits.
Large scale production and demonstration of disease
free planting material of Citrus (Nagpur mandrin) is
being done through shoot tip grafting. During the year
nearly 35,000 bud grafts were released to the farmers
and these are now being evaluated in the farmer's
field. Disease free planting stock has been raised in
the nursery for multiplication during the current year.
A multiplex diagnostic kit for detection of viruses,
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viroids and greening bacteria is being developed.
PCR detection of greening bacterium and Citrus
Yellow Mosaic Virus in Citrus tissues was
standardized using a simplified template preparation
protocol. Both the greening bacterium and CMBV
were detected through PCR when the eluted liquid
from the spotted membrane was used. The eluted
liquid was found comparable in detection efficacy to a
multi-step laboratory method or a commercial kit for
nucleic acid preparation. The protocol is simple,
inexpensive, rapid, and applicable to large-scale
survey of citrus trees. Studies are also continuing on
molecular characterization and detection of citrus
yellow mosaic virus.
A programme has been supported at IIHR, Bangalore
and CISH, Lucknow for characterization of the
germplasm of mango from Southern and Northern
region. Based on microsatellite enrichment method,
STMS primers have been developed. Using four
mango species (Mangifera odorata, M. zeylanica, M.
andamanica, M.camptosperma) and eleven cultivars
(Muvandan, Kurukkan, Alphonso, Raspuri, Totapuri,
Langra, Dashehari, Neelum, Kesar, Padari, Bhutto
Bomaby) genescan analysis was done for 30 STMS
primers with the help of automated DNA sequencer.
Many of these primers amplified all the species used
here indicating cross species compatibility. 13 ISSR
primers were used to screen the 48 accessions of
mango, among these 11 primers yielded distinct
polymorphic products, cumulatively amplifying 160
bands ISSR 5 yielded 14 polymorphic bands and 7
monomorphic bands.Based on similarity matrix,
dendogram using UPGMA tree was constructed,
which has efficiently clustered the accession from
these two regions.
Expression of Bougainvillea antiviral protein gene is
being studied at IIHR, Bangalore to develop virus
resistant tomato plants. The cDNA encoding
Bougainvillea anti viral protein was cloned into
pMALc2X expression vector. After confirmation for
the presence of the insert, the recombinant vector
was transformed into E.coli. The transformants were
selected by blue white colour. The transformants
growth was slower when induced with IPTG which is
Research and Development