NICHE AREA OF EXCELLENCE
Integrated Centre for Drought Research
Department of Crop Physiology, University of Agricultural Sciences, Bengaluru 560 065




Subprogram 1:  Genotypes with constituent drought adaptive traits

Germplasm characterization to identify donor lines with relevant drought adaptive traits

Identification of superior trait donor parents is crucial.  The focus of this sub-program is to develop suitable high throughput phenotyping strategies and identify desirable trait donor genotypes from diverse germplasm accessions.  Some input related ad-hoc projects, Center of Excellence also contributed to the outcome of this sub-program. 

Traits associated with water relations such as water mining, water conservation and superior metabolism under reduced water status such as WUE, cellular level tolerance have paramount significance. In this sub-program, donor parents possessing these adaptive drought tolerance characteristics have been identified from a large number of genotypes/breeding lines.

Root structures provide an unique option to examine the variability in root traits.  Significant variability in root traits is noticed in several crop species like sunflower, rice, groundnut, finger millet, mulberry etc. Promising trait donor parents have been identified.








Finger millet: GE 113; GPU 45 and GE 4732
Sunflower:   CMS-L-(DS) 207B, IB 66, IB 79, IB 90, IB 20
Rice:   AC35310; AC39020; IET16348; PS339; JBT 36/14.

Genetic variability in stress response is seen only when the plants are exposed to a sub-lethal induction stress. Based on this concept, temperature induction response (TIR) was developed. TIR is a high throughput technique to assess genetic variability in cellular level tolerance.







Promising donor lines for cellular level tolerance in rice have been identified:  PS-349; PS-350; Vandana; PS-400; PS-384
Stable isotope ratios are being adopted to assess variability in Water Use Efficiency in several crop species and promising lines have been identified.

Sunflower:     CMS-L-(DS) 207B, IB 13, IB 92, IB 90, IB 47, IB 86
Rice:    AC39020; JBT-37/4; IET-16348; AC-35417; AC-35310
Rice (Spikelet fertility):   AC-35633, JBT-38/26, AC-35004, PS-305

Sub-program 2:  Genes for improving drought tolerance

Transcriptional profiling and activation tagging to identify relevant candidate genes

Abiotic stress tolerant plants are known to have efficient adaptive mechanism and exploitation of such characters can be useful for crop improvement. With this background, drought specific genes are being cloned from drought hardy crop finger millet (Elucine corocana L) under NAE research programs and validated. Other related adhoc projects supported DBT, Govt. of India and in house student research programs also contributed to the outcome of this sub-project.

  • Novel gravimetric approach for drought induction developed to simulate field conditions, and different levels of soil field capacities were maintained to induce expression of drought specific gene.
  • Identified unique genes through the analysis of cDNA libraries and drought specific transcriptome, and many ESTs have been characterized and sequence information submitted to NCBI database.
  • The genes identified belongs to both upstream transcription regulatory (protein kinases, transcription factors) and downstream functional groups (enzymes and other protective proteins), indicating that libraries can be a good source for prospecting drought genes. Some of the useful genes identified are Glutaredoxin, HSP16.9, HSP17.2, HSP17.9, LEA3, Peptidyl prolyl cis trans isomerase, Translation initiation factor5A, transcription factors like NAM, bZIP, MYC and GBF.   
  • A few drought genes have been functionally characterized by down regulation studies (RNAi and VIGS) in model systems.
  • A few full-length transcription factors (EcNAC1, EcNAM1, EcGBF1, EcZnfinger, EcMYC, and EcbZIP) cloned are being evaluated by over expression studies.
  • Suitable over expression cassettes for over expression of specific validated genes (P5CS, mtld, Helicases, ipt, DREBs, NAC, Annexin and HSF4) have been made, and  used for developing transgenic crop plants to improve drought resistance. 
  • Developed approaches to stack genes with stress specific inducible promoters and over expression cassettes carrying multiple genes are being evaluated. 
     











Subprogram 3:  Transgenics for improving the drought tolerance

Development of transgenics expressing stress genes and their evaluation

Trait pyramiding can be effectively achieved through introducing novel validated genes through transgenic technology.  Towards this end, robust and reproducible transformation protocols has been developed using specific recipient lines in groundnut, rice and sunflower.  The major focus of this sub programme has been to develop transformants expressing upstream regulatory genes associated with stress tolerance in identified genotypes with superior water relations traits. With this approach, transgenics developed in groundnut were productive under stress confirming the proof of concept that trait pyramiding by transgenics is promising.

In planta transformation protocol has been developed in groundnut, rice and suflower. The protocol involves in planta inoculation of embryo axes of germinating seeds and allowing them to grow into seedlings ex vitro. The primary transformants being chimeric, emphasis has been on herbicide selection to identify the putative transformants in T1 generation. In house student programmes also contributed to the outcome of this component.

 



Groundnut transgenics with improved stress adaptation

Transgenics developed expressing Helicases and DREBs (DREB1A, 1B and 2A) in the background of genotypes with superior water relations are promising. Many promising events in advanced generations showed higher WUE and were tolerant to moisture stress besides being productive. COE, DBT also contributed to this sub-programme with collaboration from ICGEB – Dr. Tuteja and Dr. M. K. Reddy,





 Groundnut transgenics expressing ipt

The plant hormone cytokinin regulates leaf senescence and sink capacity in plants. Transgenic plants over expressing ipt a rate limiting enzyme in cytokinin synthesis were developed. Some transgenic events showed reduced leaf senescence and increased pod size. Chlorophyll stability on exposure to simulated stress was also high in transformants
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Transgenics have also been developed with other upstream regulatory genes like Hsf and NACs. Evaluation is under progress for the identification of promising events.

With a similar concept to pyramid traits, superior trait donor parents with higher water mining and water use efficiency were identified in sunflower and rice. Transgenics are being developed using upstream regulatory genes.

Subprogram 4:  Trait introgression to improve drought tolerance through molecular breeding

a)Identification of QTL for specific drought traits
b)Development of double haploid as a tool to
    i) Introgress traits
   ii) Develop mapping populations

Several polygenic traits that have relevance to drought tolerance need to be pyramided onto a single elite genetic background.  Thus, mapping QTL regions associated with drought adaptive traits and exploiting the markers for trait introgression through Molecular breeding is the major emphasis of this sub-program. The goal is to improve the adaptation and productivity of rice grown under semi irrigated aerobic conditions. Aerobic cultivation is a potential option to save substantial amount of irrigated water.

Two QTL mapping strategies are being adopted for the identification of DNA based markers linked with relevant traits such as WUE, root traits and other related traits like spike let sterility.

Bi-parental QTL mapping

A trait specific mapping population segregating for WUE was developed by crossing Thanu (low Δ13C) and IET-15963 (high Δ13C) and has been advanced to F7 towards generation of RIL population. A few of these segregants have been identified as promising lines and are being evaluated under aerobic field trial along with other checks.



Frequency distribution of Δ13C

 
Molecular and phenotypic characterization of the population led to identification of a few major QTLs governing WUE and other related parameters and their chromosomal location has been mapped.





Linkage disequilibrium based association mapping

Frequency distribution for Root weight and Δ13C


Phenotyping the diverse germplasms accession for various physiological traits like water mining, water conservation, intrinsic tolerance and spikelet fertility are being carried out.
 





A More robust strategy of linkage disequilibrium (LD) based association mapping has been adopted. Based on the genetic diversity determined by using microsatellite markers, the population structure has been assessed. LD decay on rice genome and marker trait association by LD mapping is being carried out.






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