Environmental stresses are serious threat to agriculture and the primary cause of crop loss worldwide and become a major challenge in our quest to achieve sustainable food production. Genetic engineering thus provides alternatives to combat this problem addressed.

1. Feb
2017
Isolation and characterization of stress inducible promoters from
Pennisetum glaucum and their role in abiotic stress adaptation
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Divya K1
, Reddy PS1
*, Bhatnagar-Mathur P1
, Reddy MK2
, Vadez V1
, Sharma KK 1
*E-mail: p.sudhakarreddy@cgiar.org
Introduction
• Environmental stresses are serious threat to agriculture and the primary cause
of crop loss worldwide and become a major challenge in our quest to achieve
sustainable food production.
• Genetic engineering thus provides alternatives to combat this problem
addressed.
• Transgenic plants developed with a strong constitutive expression of functional
genes and/or transcription factors often suffer from undesirable phenotypes.
• It is therefore desirable to generate transgenic plants that accumulate
transgenic products only under stress conditions.
• Use of stress inducible promoters has proven to be an alternative strategy for
elimination of such phenotypes induced by constitutive transgene expression.
Objective: Development of the abiotic stress tolerant crops
The relative expression of five abiotic stress inducible genes under different
abiotic stress conditions. The fold change between 2 to 5 fold are given a single
(+) or (-) sign for up regulation and down regulation respectively. Similarly (++) or
(- -) denotes 5 to 10 fold change, (+++) or (- - -) denotes 10 to 15 and (++++) or (-
- - -) denotes 15 fold or above change in the expression values.
Heat (45 o
C) Cold (4 o
C) Drought (withholding the water for 48 h) Salt (250 mM)
Hsp10 ++++ - + +
Hsc70 ++++ - ++ +/-
Lea +++ ++++ + ++++
Dhn + - + ++++
APX ++++ ++ + +
Isolation and cis-motif analysis of promoter regions of five abiotic stress inducible genes.
Schematic diagram of pMDC164 vector
Cloning in to pMDC164 vector
Hsp10S Dhn Hsc70 Apx Lea
M 1 2 3 4 5 6 7 8 TC TC
1 2 3 4 5 6 7 8 M TC TC P N
Cloning, transformation and
molecular confirmation of the
tobacco transgenic plants.
Conclusion
• PgHsc70 pro and PgLea pro were active in the heat, PgDhn pro in heat, cold and
drought and PgApx pro in drought stress conditions.
• PgHsc70 pro and PgDhn pro can be used to drive the expression of selected
target gene to develop abiotic stress tolerant crops.
References
• Reddy PS, Mahanty S, Kaul T, Nair S, Sopory SK, Reddy MK (2008) A high-
throughput genome-walking method and its use for cloning unknown flanking
sequences. Anal Biochem 381:248-253.
Acknowledgments
Innovation in Science Pursuit for Inspired Research (INSPIRE), Department of
Science and Technology, Govt. of India for the research grant through INSPIRE
Faculty Program (Award No. IFA-LSPA-06). This work has been undertaken as part
of the CGIAR Research Program on Dryland Cereals.
GUS Histochemical assay. Symbols, + (mild expression), ++ (strong expression) and -
(no expression) represents the intensity of GUS activity.
2
International Centre for Genetic Engineering and Biotechnology (ICGEB), Aruna Asaf Ali Marg, New Delhi-110 067, India
1
International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Hyderabad, 502 324, Telangana, India
Results
Hsp10 Hsc70 Apx
LeaRab
Walker
Primers
Phi29 DNA
polymerase
Genomic DNA
Whole genome is
copied into large
number of
overlapping
fragments with
unique sequence
Cis-elements of stress inducible promoters
ABREHSE LTR
GC-motif CircadianSkn-1_motif
ARE
CGTCA
TGACGTGABox-W1MBS
CCGTCC-box GCN4_motif
-100 -300-200 -400 -600-500 -700 -800-1
APX
Hsc70
Dhn
Lea
Hsp10
-900
Reddy et al., 2008
1. Expression profiling of five abiotic stress inducible genes.
2. Isolation and analysis of promoter regions of five abiotic stress inducible genes.
3. Construction of plant expression vector with five different promoters.
Relative GUS mRNA level in
abiotic stressed transgenic
tobacco plants.
4. Functional validation in tobacco using histochemical assay and molecular approaches.
Drought tolerant crop
Pearl millet
Crop plants Model plants
Rice
Legumes
Sorghum
MaizeWheat
Evaluation in model plants
Stress tolerant crops
Agriculturally important crops