ANNUAL REPORT - Department of Biotechnology

Signal Transduction
In a study supported at JNU to study signal
transduction machineries in plant under osmotic
stress, complete physiological characterization of
genotypes of Brassica campestris, B. nigra,
B.oleracea, B. juncea, B. napus and B. carinata for
their tolerance has been achieved under laboratory
conditions. Primers have been designed to isolate
the middle portion of the hybrid type Histidine kinase
from B. juncea (CS52) which has resulted in cloning
and sequencing of G80 bp fragment. Novel
transcription factors have also been cloned from
three B. juncea varieties specific to salinity tolerance.
RNA has been isolated and primers were designed
using Atmyb2 conserved domain. The PCR-product
was amplified and cDNA is being cloned. An efficient
plant regeneration and transformation system has
been optimized in selected commercial cultivars of B.
juncea. A study has been supported at ICGEB to
clone and characterize stress responsive promoters
from Brassica sps. Putative promoter regions for
stress related genes like SoS2, Histidine kinase
(HKI), Myb and MyC like factors have been cloned.
The amplified fragments for putative promoters have
been cloned, which show 50-80% homology. Further
work is going on to assess the true stress inducibility
of these promoter sequences.
Scientists at NCPGR, New Delhi has been able to
clone 8 members of mitogen activated protein kinase
family (MAPK) from rice. From the 8 clones, four full
length genes have been obtained. They have further
been cloned in PGEX vector having GST protein
fusion for heterologous expression in bacterial
system. MAP kinase induced by heat has been
identified and work is on to transform rice with
different constructs.To understand the mechanism
operative during early signaling events mediating
auxin dependent abiotic stress tolerance in
groundnut at Calcutta University, CDPK has been
characterized from Arachis hypogea showing 88%
homology to a stress responsive CDPK from
Arabidopsis. Result indicated that Dephsphorylation
of AhCPK2 is not a response to water loss per se but
to a change of water potential. No such change was
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noted in the presence of ABA treatments indicating
the noted oscillation of phosphorylation of this kinase
to be an upstream event.
Based on in silico analysis, 46-stress responsive
transcription factors were identified from Arabidopsis
highly stress responsive functional genes have also
been identified and their promoter analyzed. No
significant difference was observed for presence of
transcription binding sites of stress responsive
transcription factors belonging to 10 families.
Functional involvement of a light signaling
component (ZBF1) in root development in
Arabidopsis. ZBF1 (Z-box binding factor 1), which
codes for a bHLH protein has recently been cloned at
NCPGR, New Delhi. The function of A novel
transcription factor of light signaling on root
development has been investigated. Mutations in
ZBF1 results in elongated roots as compared to wild
type plants.
At NBRI, the expression of three Ethylene
Responsive Factors (ERFs) was found ten days after
watering was stopped. Increased transcript
accumulation was also observed after a cold shock
of 4 hours for LeERF5 and LeERF6 while expression
of all three ERFs was repressed after a heat shock at
42°C for one hour. In order to functionally analyse
these genes, they were expressed in a bacterial
expression system in the vector pET28. The proteins
isolated are currently being analysed. Transgenic
tomato plants that overexpress the three ERFs have
also been raised and are being studied for
morphological and physiological changes.
Floral Development
To understand how floral organ development and
regulation of size & shape of floral organs and leaves
take place at molecular level, studies have been
conducted at IISc, Bangalore on Arabidopsis.
17,000 EMS-mutagenized M2-seeds were screened
and based on phenotypes, mutants have been
categorized. To check the DNA binding properties of
wild type TCPY, consensus binding site of TCP4 was
determined using random binding site selection
Research and Development