On Monday, June 1st, SIVB held an oral presentation competition for students. After their presentation, the students answer questions from the audience and the judges. The judges, Valerie Pence, Michael Kane and Randall Niedz, chose 3 finalists and ranked, Steven Samuels from Tuskegee University for 1st prize, Jose Tovar from Arkansas State University for 2nd prize and Kerri Neugebauer from Kanas State University for 3rd price. Plaques and cash awards were presented at the Plant Section Business Meeting. We encourage all qualified students to enter their research for the 2016 Plant Biotechnology oral presentation.
Submitted by Cecilia Zapata
In-Planta Expression of Synthetic Lytic Peptides for the Potential Inhibition of Human Virus Replication
Recombinant plant systems offer economic alternatives to producing large amounts of pharmaceutical proteins as well as providing the most promising opportunity to supply low-cost drugs and vaccines for major diseases such as HIV. Treatments of infectious diseases in humans and animals have traditionally been targeted by chemically synthesized drugs, with the majority of the burden of cost falling on the individual in need of treatment. With the new revolution of producing therapeutic compounds, such as peptides in plant based systems, the cost of production is dramatically decreased. Synthetic antiviral peptides capable of inhibiting the progression of HIV have been developed at Tuskegee University. These synthetic lytic peptides genes were designed with an intron to facilitate cloning in bacteria and were mobilized in Agrobacterium tumefaciens strain EHA105 for transformation of sweetpotato cultivar PI318846-3. Seven transgenic plants were confirmed by Southern and RT-PCR analyses, leading to testing of the efficacy and toxicity of crude and purified sweetpotato extracts. Treatment of infected cells showed an 80% inhibition of DH12 and SF162, two well characterized pseudovirus strains, in neutralization assays. Peptide treated DH12 and SF162 show significant reduction as compare to no peptide control. Further biosafety, nutritional, and efficacy tests will determine agronomic performance, proximate quality and potency of transgenics respectively. Successful development of sweetpotato expressing this novel therapeutic compound can be both a powerful tool in treatment of the HIV epidemic, as well as a road map for future treatment of viral mediated diseases. Work supported by Tuskegee University GWCAES, NIFA-Evans-Allen, Project Export, and UC-Davis-TU CREATE Igert, Iowa State University GWCIP.
Steven Samuels, CAENS Department of Agriculture & Environmental Sciences, Plant Biotech & Genomics Research Lab, Tuskegee University, Tuskegee AL 36088. In Vitro Cellular and Developmental Biology, 51:S33-S34, 2015
Basis for Engineering an Improved Processing Trait in Sugar Beets
We are investigating a novel biotechnical application of a thermally-tolerant pectin methylesterase (TT-PME) for improving energy efficiency in sugar beet processing. Sucrose diffusion from beet root slices occurs in 60-65°C water, optimal conditions for TT-PME activity. Separating water from wet pulp is necessary to stabilize it for storage and reducing weight for shipping. Drying pulp is a highly energy intensive process (consumes up to 30% of total energy in a processing factory). Beet pulp’s high water binding is due to its high content of pectin, a complex cell wall polysaccharide that entraps water. We hypothesize TT-PME action can reduce water binding in pulp by promoting calcium-mediated crosslinking, which will facilitate improved mechanical de-watering, resulting in lower energy cost and environmental footprint for drying pulp. To test this hypothesis, we developed an in vitro assay to quantify water binding in pulp. We found that TT-PME action reduced water binding in beet pulp by 27% with supplemented calcium. This evidence for functional benefit provides basis for our goal to develop transgenic beet roots that express TT-PME as an improved output trait. To support this goal, we developed peptide-based monospecific antibodies to detect TT-PME in transgenic plant tissues. Current experiments are directed to assess the function and impact of the pro-peptide domain in expressing active recombinant enzyme. For this, we are using the rapid and well-established Nicotiana benthamiana transient expression system. This presentation will summarize our findings on TT-PME’s pulp water binding reduction, specific antibody production, and progress on N. benthamiana TT-PME expression.
Jose C. Tovar, Molecular Biosciences Graduate Program and Arkansas Biosciences Institute, Arkansas State University, Jonesboro, AK. In Vitro Cellular and Developmental Biology, 51:S32-S33, 2015
Wheat Gene Expression Differences Induced by Six Races of Puccinia triticina
Puccinia triticina, the casual agent of wheat leaf rust, is a devastating disease that can cause up to 40% yield loss. During fungal infection the host plant recognizes pathogen effectors, which trigger a host defense response. Changes in the pathogen effectors due to host selection pressure are responsible for the rapid development of new rust races and make durable resistance hard to obtain. The objectives of this study are to identify and characterize wheat genes that are utilized by races differently throughout infection and to understand functions of these genes using gene silencing. Six races of leaf rust were inoculated on a susceptible wheat variety and tissue was collected at six days post inoculation. RNA was sequenced and 63 wheat genes were identified that showed varying expression in response to the six races. 54 of these genes were evaluated in a time course study from zero days to six days post inoculation with the same six races. Real-time PCR was then used to analyze the timing of expression during early infection. The characterized genes have proposed functions involved in plant defense and stress, energy and metabolism, protein transport, replication, and RNA binding. Majority of the candidate genes showed three main expression patterns. However, race specific expression was found in three wheat genes that are affected by race shifts on Lr2A, Lr2C, and Lr17A. Sixteen potential susceptibility genes were also identified. Host susceptibility genes could be altered to provide durable resistance. RNAi was used to silence seven wheat genes to further understand their roles in leaf rust infection. T0 and T1 plants have been obtained and confirmed for the gene of interest. T2 plants were inoculated and observed for changes in susceptibility.
Kerri Neugebauer, Kansas State University, Department of Plant Pathology, 4024 Throckmorton, Manhattan, KS 66506. In Vitro Cellular and Developmental Biology, 51:S32, 2015