Progress 01/01/12 to 12/31/12
Outputs
OUTPUTS: Food security is an increasing global concern. Yam (Dioscorea spp.) is one of the most important staple food crops for over 400 million people. Yam tubers contain many secondary metabolites known to be useful to human health. Yam starch is suitable for many industrial applications. Yet, yam is underutilized and is lagging behind the modern genomic era. This project is designed to contribute to the general effort to accelerate improvement of yam production, industrial utilization, and food security. The project activities specifically include: 1) to develop of a tissue culture technique to efficiently transfer foreign genes to yam via Agrobacterium tumefaciens, 2) to provide a source of genes for comprehensive understanding of the regulatory networks governing certain traits in yams, 3) to provide genomic data that can be used in high-throughput approaches in yam quality enhancement via breeding and/or genetic engineering, 4) to increase the number of yam nucleotide sequences in the Genebank and, 5) to train and prepare minority graduate students for successful career in science and/or academia. PARTICIPANTS: Dr. Ramesh Kantety Part of this project was built on the strength and the expertise of the co-PI, Dr. Kantety in gene expression studies including sequencing and analysis of expressed sequence tags (ESTs). Unfortunately, Dr. Kantety became unavailable due to sickness which kept him out of research activities. He passed in summer 2012. Therefore, cDNA library screening, sequencing and EST analysis are yet to be completed TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
In this Evans Allen project, work was initiated to induce callus in edible yam. Successful callus induction was obtained using petiole explants from in vitro yam plants cultured on MS medium, supplemented with 3 percent sucrose, 1.5 millimolar putrescine and naphtoxy acetic acid (NOA) at 20-50 micro molar. Callus initiation was initiated within 8 to 12 weeks, and less than 40 percent explants responded. Attempts to induce callus from leaf, stem and root tissues failed despite the application of a wide range of hormonal and physical treatments including auxins, cytokinins, dark and light. Although the three genotypes tested (D. alata cv Bete Bete, D. rotundata cv Kponan Fissa and D. cayenensis cv Krengle IB14) responded well to callus induction, Bete Bete was the least responsive. Yam callus was successfully propagated by omitting putrescine from the MS medium, and by supplementing the medium with either NOA (20 and 50 micro molar), or 20 and 50 micro molar naphthalene acetic acid (NAA). NAA at 20 micro molar produced the healthiest and good growing callus. Regardless the hormone type and the concentration, root-like structures developed on the majority of yam callus explants, demonstrating the organogenic potentials of yam callus. Therefore, culture conditions are being optimized to regenerate shoots and/or embryos from yam callus. In vitro yam plantlets from D. alata cv. Bete bete and D. cayenensis cv. IB14 were acclimatized and transferred in an Enconair A60 growth chamber set at 32 degree Celcius/24 degree celcius and 14h/8h day/night. Yam plantlets were subjected to drought and salt stresses. Drought stress was applied by withholding watering of yam plants while the control plants were watered once every three days. After about 15 days, plants under drought treatments started losing their turgescence. Root tips and young leaves were collected for total RNA extraction. Salt stress was performed by watering the plants with 100 millimolar NaCl solution. Plants were watered once every three days. After 30 days, there was no significant morphological difference between the non-treated and salt treated plants. Root tips and young leaves were collected, and total RNA was extracted. Similarly yam callus was subjected to salt and drought stresses by subculture on medium supplemented with NaCl (10 and 100 millimolar) and mannitol (30 and 300 mM) respectively. Total RNA was extracted from yam root and callus. A total of 13 cDNA libraries have been constructed from tissues of D. cayenensis cv IB14, and D. alata cv Bete bete. Libraries were constructed using the SMARTTM technology (Clontech laboratories Inc. Mountain View, CA), arrayed in 96 well plates and stored at -80 degree Celcius. Library screening, DNA sequencing and analysis of gene expression are yet to be performed. This project has graduated two female African American students for Master degree in Food Science, and undergraduate minority students were trained in plant biotechnology techniques including plant tissue culture and plant propagation, molecular biology (extraction and purification plant DNA and bacterial plasmids, PCR, protein extraction, SDS PAGE and ELISA)
Publications
- Two Master Thesis were produced Charvais Bogan (2011) Enhancement of Agrobacterium-mediated genetic transformation of the tuber crop yam (Dioscorea spp). Master Thesis Alabama A&M University Alishia Bush (2012) In Vitro Induction of Callus from the Tuber Crop Yam (Dioscorea Spp.). Master Thesis Alabama A&M University
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