Progress 04/15/24 to 04/14/25
Outputs
Target Audience:Undergraduate and graduate students, plant biology researchers in academia, and scientists in biotechnology industries focus on oilseed improvement, photosynthesis, and renewable energy. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?The project provided a training opportunity for undergraduate and graduate students during the reporting period. How have the results been disseminated to communities of interest?Publications Deo, B., Muthan, B., Cruise, T.L.A., Mukherjee, T., Allen, D.K., & Sanjaya, S.A. (2025). Flue Gas Desulfurized Gypsum as a Sustainable Amendment for Coal Mine Soil Reclamation and Bioenergy Crop Production. Science of the Total Environment (In Review). Sanjaya, S. (2025). How biotechnology can revolutionize agriculture: feeding the world and powering the future in the face of climate change. In proceedings of an International Conference on Future perspectives in plant science (ICFPPS-2025). PSG College of Arts and Science, Coimbatore, India. pp-24. OSBN 978-81-980915-0-5. Poster Presentations: Muthan, B., Khatiwada, S., and Sanjaya, S. (2024). Application of Gene Technology for Increasing Oil in Bioenergy Crops. Midwest ASPB meeting, Purdue University, March 2024. Muthan, B., Khatiwada, S., Jayaraman, K and Sanju Sanjaya (2024). Genetic engineering of switchgrass biomass for bioenergy production. ASPB meeting, Honolulu, Hawaii, June 2024. Bagyalakshmi Muthan, Jie Wang, Ruth Welti, Dylan K. Kosma, Linhui Yu, Bikash Deo, Subhiksha Khatiwada, Vijaya K.R. Vulavala, Kevin L. Childs, Changcheng Xu, Timothy P. Durrett, Sanju A. Sanjaya (2024). How Aquatic Plants Develop Tolerance to Heavy Metal Toxicity. 26th International Symposium On Plant Lipids, UNL, July 2024. Oral Presentations: Sanjaya, S. (2024). Understanding How Plants Tolerate Heavy Metal Stress: The molecular Mechanisms. East Tennessee University, TN, USA. Sanjaya, S. (2025). Lipid metabolism in duckweed under heavy metal stress conditions. Gordon Research Conference on Plant Lipids: Structure, metabolism and functions, Pomona, CA, USA. Sanjaya, S. (2025). Molecular mechanisms of heavy metal stress tolerance in Spirodela polyrhiza. Plant and Animal Genome Conference 32 (PAG32), San Diego, CA, USA. Sanjaya, S. (2025). Molecular Mechanisms of Plant Tolerance to Heavy Metal Stress. North Carolina A & T State University, Greensboro, NC, USA. Sanjaya, S. (2025). Genetic Transformation of Recalcitrant Upland Energy Crop Switchgrass for Enhanced Oil Yield. Plant and Animal Genome Conference, Asia, New Delhi, India. Sanjaya, S. (2025). How biotechnology can revolutionize agriculture: feeding the world and powering the future in the face of climate change. PSG College of Arts and Science, Coimbatore, India. Sanjaya, S. (2025). Potentials of Biotechnology to Produce Renewable Hydrocarbons in Plants for Dealing with the Effect of Climate Change. University of Mysore, India. Sanjaya, S. (2025). Metabolic engineering of hydrocarbons in plants for bioenergy production. PES University, Bangalore, India Sanjaya, S. (2025). Can the application of biotechnology techniques help to feed and fuel the world? M.S. Ramaiah College of Arts, Science & Commerce, Bangalore, India. What do you plan to do during the next reporting period to accomplish the goals?We will continue investigating the biochemical and molecular characteristics of transgenic switchgrass. Comprehensive analyses of key primary metabolites and seed storage compounds will be conducted in both T? and T? transgenic lines. Concurrently, seed bulking is underway to support forthcoming performance evaluations under controlled-environment and greenhouse conditions. Additional efforts will include molecular genotyping of transgenic events and detailed assessments of growth and yield parameters, including photosynthetic efficiency and key agronomic traits.
Impacts
What was accomplished under these goals?
A robust and reproducible protocol was established for the surface sterilization and in vitro callus induction of mature seeds of Panicum virgatum L. (switchgrass), cultivar Alamo. Approximately 6-7 g of mature seeds were soaked in 100 mL of sterile distilled water in a 250 mL conical flask and agitated on a rotary shaker at 200 rpm for 30 minutes at room temperature. After decantation, seeds were rinsed 3-4 times with sterile distilled water to remove debris. Surface sterilization was achieved by immersing the seeds in 100 mL of full-strength commercial Clorox® bleach with gentle stirring for 2 hours, followed by 4-5 rinses with sterile distilled water to remove residual bleach. Sterilized and dehusked seeds were transferred to callus induction medium (CIM) and incubated in the dark at 28°C for 3-4 weeks, resulting in efficient sterilization and robust callus formation. Murashige and Skoog (MS) medium was selected as the CIM due to its demonstrated effectiveness in promoting the formation of highly friable and regenerable calluses in switchgrass (cv. Alamo). To maintain optimal callus proliferation, the medium was refreshed biweekly over a 6-8 week culture period. Actively dividing calluses were used as explants for Agrobacterium tumefaciens-mediated transformation. Calluses approximately 0.5-1.0 cm in diameter were pre-cultured on fresh CIM for 3 days before being immersed in Agrobacterium suspensions carrying target plasmid constructs. The explants underwent vacuum infiltration, gentle agitation, and a 2-day desiccation period, followed by a 2-day rest on callus resting medium. They were then transferred to selective CIM containing 50 mg/L Hygromycin B for two rounds of selection. Hygromycin-resistant calluses were moved to regeneration medium (REG) supplemented with 15 mg/L Hygromycin B. Successful regeneration was characterized by red-purple pigmentation, the formation of green nodular tissues, and the emergence of visible shoot meristems. Shoots reaching 1-2 cm in height were transferred to half-strength MS medium for rooting. Fully rooted plantlets were acclimatized in soil pots and maintained under controlled growth chamber or greenhouse conditions. All putative transgenic lines--including those containing single and multi-gene constructs--were advanced to produce T? seeds. These lines are currently being propagated for subsequent molecular and biochemical characterization to validate transgene integration and expression.
Publications
- Type:
Other Journal Articles
Status:
Under Review
Year Published:
2025
Citation:
Deo, B., Muthan, B., Cruise, T.L.A., Mukherjee, T., Allen, D.K., & Sanjaya, S.A. (2025). Flue Gas Desulfurized Gypsum as a Sustainable Amendment for Coal Mine Soil Reclamation and Bioenergy Crop Production. Science of the Total Environment (In Review).
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2025
Citation:
Sanjaya, S. (2025). How biotechnology can revolutionize agriculture: feeding the world and powering the future in the face of climate change. In proceedings of an International Conference on Future perspectives in plant science (ICFPPS-2025). PSG College of Arts and Science, Coimbatore, India. pp-24. OSBN 978-81-980915-0-5.
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Progress 04/15/23 to 04/14/24
Outputs
Target Audience:The research participants included undergraduate and graduate students, visiting scholars, plant biology researchers in academia, scientists in biotechnology industries focused on bioenergy, surface coal mine land reclamation agencies, farmers, local non-governmental organizations, and farmers. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?Through this project, we have provided experiential learning opportunities for undergraduate and graduate students, visiting scholars at WVSU, and researchers in academia and industry. Project outcomes were shared both formally and informally with scientists focused on agriculture, bioenergy, plant biology, renewable energy, farmers, and post-coal mining reclamation. How have the results been disseminated to communities of interest?Poster presentations: Muthan, B., Khatiwada, S., and Sanjaya, S. (2024). Application of Gene Technology for Increasing Oil in Bioenergy Crops. Midwest ASPB meeting, Purdue University, 2024. Muthan, B., Khatiwada, S., Jayaraman, K and Sanju Sanjaya (2024). Increasing energy density of switchgrass biomass for bioenergy production. In the 21st Research Symposium of the Association of 1890 Research Directors. Nashville, TN, April 2024. Cole, G., Patil, S., Muthan, B., and Sanjaya, S. (2024). Understanding the Effect of Carbohydrates on Seed Germination and Establishment in Arabidopsis. In the 21st Research Symposium of the Association of 1890 Research Directors. Nashville, TN, April 2024. Oral presentations: Sanjaya, S. (2024). Climate Change and Biotechnology for Improving Production of Food and Fuels. In the 21st Research Symposium of the Association of 1890 Research Directors. Nashville, TN, April 2024. Sanjaya. Molecular mechanisms of heavy metal stress tolerance in plants. Kansas State University, February 2024. Sanjaya. Can we meet the world's growing demand for food and energy using biotechnology, University of Tennessee, April 2023. Sanjaya. Potentials of Biotechnology to Produce Renewable Hydrocarbons in Plants for Dealing with the Effect of Climate Change. New frontiers of natural sciences: 2nd international selçuk meeting October 02-04, 2023 Konya-Türkiye Sanjay. Energy and environmental science research and educational opportunities and challenges in West Virginia, University of Tennessee, October 2023. What do you plan to do during the next reporting period to accomplish the goals?We will continue to study the effects of environmental resilience in switchgrass using biochemical and genomics experiments. We will establish robust Agrobacterium-mediated transformation systems to introduce target genes into switchgrass and generate many transgenic plants expressing these genes. Additionally, we will develop genetic, biochemical, and agronomic methods to select for switchgrass lines with increased biomass.
Impacts
What was accomplished under these goals?
The best upland and lowland switchgrass varieties were selected for further analysis based on their growth and physiological performance in response to coal mine soil conditions. These varieties were subjected to environmental stresses including soil type, high temperature, drought, and solar radiation. Studies investigating the physiological and biochemical responses of switchgrass to these stresses are in progress. Using gene-specific primers, we have cloned target genes individually and in combination into binary vectors containing monocot-specific promoters and plant selection marker genes. We then mobilized the binary vectors into Agrobacterium and confirmed their presence using PCR with gene-specific primers. Callus cultures free of microbes have been successfully developed from switchgrass seeds, and we have optimized the antibiotic and plant growth regulator combinations to efficiently transform and regenerate transgenic plants. A few transgenic plantlets containing single or double target genes have been generated through prolonged antibiotic selection. Work is underway to regenerate sufficient numbers of transgenic plants and perform genetic analysis.
Publications
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2024
Citation:
Sanjaya, S. (2024). Climate Change and Biotechnology for Improving the Production of Food and Fuels. Pp 265. In the 21st Research Symposium of the Association of 1890 Research Directors. Nashville, TN, April 2024.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2024
Citation:
Muthan, B., Khatiwada, S., Jayaraman, K and Sanju Sanjaya (2024). Increasing energy density of switchgrass biomass for bioenergy production. Pp 521. In the 21st Research Symposium of the Association of 1890 Research Directors. Nashville, TN, April 2024.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2024
Citation:
Cole, G., Patil, S., Muthan, B., and Sanjaya, S. (2024). Understanding the Effect of Carbohydrates on Seed Germination and Establishment in Arabidopsis. Pp 429-430. In the 21st Research Symposium of the Association of 1890 Research Directors. Nashville, TN, April 2024.
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