The following student awards were presented at the 2012 World Congress on In Vitro Biology in Bellevue, Washington. Information on additional awardees at the 2012 World Congress was presented in the July – September issue of the In Vitro Report Information related to the available specific student awards can be found on the SIVB website (www.sivb.net) or by contacting the SIVB Business Office at (919) 562-0600, [email protected], or Dr. Pamela Weathers, Chair, Student Affairs and Awards Committee, at [email protected].
2012 HOPE E. HOPPS AWARD AND 2012 SIVB STUDENT TRAVEL AWARD
Development of a Genetic Transformation Protocol for Interspecific Hybrids of Elephantgrass (Pennisetum purpureum) x Pearl Millet (Pennisetum glaucum)
Elizabeth Mayers
Interspecific hybrids of elephantgrass (Pennisetum purpureum) x pearl millet (Pennisetum glaucum) are among the most productive perennial grasses for biofuel and forage production in subtropical and tropical regions. Elephantgrass produces wind dispersed seeds which may contribute to invasiveness. In contrast, triploid, interspecific hybrids of elephantgrass x pearl millet are completely sterile and seedless which will enhance biocontainment. We recently generated and selected triploid, sterile, interspecific hybrids with superior biomass yields and persistence. A genetic transformation protocol for further improvement of interspecific hybrids of elephantgrass x pearl millet was lacking and its development was our primary objective. This involves the optimization of tissue culture, gene transfer and selection parameters. Five breeding lines with superior biomass yield and persistence were evaluated for tissue culture response. All of the lines were able to induce regenerable callus with line PMN 51 regenerating the most shoots per explant and callus. This line was therefore selected for biolistic gene transfer with selectable marker (nptII) constructs differing in the constitutive promoter (enhanced 35S or ubiquitin). The timing of selection as well as concentration and type of the selective agent (paromomycin or geneticin) were also compared. Transgenic nature of the regenerated plants was confirmed by PCR and NPTII-immuno- chromatography. Data from Southern blot analysis and NPTII-ELISA will also be presented. This is the first report of transgenic interspecific hybrids of elephantgrass x pearl millet. This genetic transformation protocol will allow the introduction of transgenes to enhance biomass production and stress tolerance while providing elevated biocontainment through the sterile, seedless nature of this promising biofuel feedstock.
Elizabeth Mayers, University of Florida, IFAS, Agronomy Dept., 3062 McCarty Hall D, McCarty Dr., Gainesville, FL 32611. In Vitro Cellular and Developmental Biology, 48:S80-81, 2012
2012 CELLULAR TOXICOLOGY AWARD
Establishing and Characterizing an Endothelial Cell Line from the Bulbus Arteriosus of Juvenile Walleye, Sander vitreus, for Physiology, Toxicology and Virology Research
Nguyen T. K. Vo
As few fish endothelial cell culture models are available, the goal was to develop a teleost endothelial cell line with properties that would allow its use for a wide number of purposes, befitting the critical roles of endothelial cells in tissue homeostasis, toxicology and infectious diseases. Cell cultures were sought from the bulbus arteriosus (BA) of walleye (WE), which is one of North America’s most important commercial and recreational freshwater fish. The BA is between the ventricle and ventral aorta of the teleost circulatory system and has an inner wall of endothelial cells. Unlike other potential sources of fish endothelial cells, the BA was large, visually distinct, and excised cleanly, permitting the easy preparation of explants. From these developed a cell line, WEBA1F. WEBA1F was maintained for at least 20 passages in L-15 medium with 10% fetal bovine serum at 26°C. The cells had cobblestone morphologies at confluency and formed capillary-like tubes in superconfluent cultures. As one physiological function of some fish endothelial cells is to ‘cleanse’ blood, WEBA1F were examined for scavenger activities. The cells highly phagocytized polystyrene beads, took up acetylated low-density lipoproteins, and responded to the scavenger receptor ligand, polyinosinic: polycytidylic acid. As the endothelium in several vertebrates is the target for dioxins, inducing CYP1A1 and endothelial dysfunction, WEBAIF were exposed to a wide range of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) concentrations. Ethoxyresorufin-O-deethylase (EROD) activity, which is a catalytic measure of CYP1A, was induced in a dose-dependent manner. Viral hemorrhagic septicemia virus (VHSV) genotype IVb has recently been detected in Great Lakes fish and appears to replicate in endothelial cells and impair their functions. Therefore WEBA1F cultures were inoculated with VHSV IVb. WEBA1F underwent cytopathic effect (CPE) and produced VHSV IVb. In summary WEBA1F is a novel endothelial cell line that should be a useful tool in studying fish endothelial physiology, toxicology and virology.
Nguyen T. K. Vo, Department of Biology, University of Waterloo, Waterloo, ON, CANADA N2L 3C5. In Vitro Cellular and Developmental Biology, 48:S32-33, 2012
2012 JOSEPH F. MORGAN AWARD AND 2012 SIVB STUDENT TRAVEL AWARD
Characterization of Bovine Herpesvirus Type 1 as a Novel Oncolytic Virus
Breanne Cuddington
Oncolytic viruses are an attractive avenue of cancer therapy due to the absence of toxic side effects often seen in current treatment modalities. HSV-1 has been studied extensively as an oncolytic virus due to the many advantages it possesses for use in virotherapy; however, there are also a number of disadvantages to this approach. To this end, we are interested in evaluating BHV-1 as an oncolytic herpesvirus. A preliminary screen measuring both virus replication and cellular viability showed that BHV-1 is cytotoxic to various human immortalized and transformed cell lines, while being relatively restricted in normal primary cells in vitro. In addition, the NCI60 panel of human cancer cell lines has been screened with BHV-1. Results of the screen show that 94% of tumour cell lines were permissive to BHV-1 with 79% showing a decrease in cellular viability. These results suggest that BHV-1 holds promise as a broad spectrum oncolytic vector that is able to infect tumour cells from a variety of histological origins. In contrast to many oncolytic viruses, cellular sensitivity does not correlate with type I IFN signaling. Furthermore, non-permissive cell lines are susceptible to infection with wild type HSV-1, which utilizes the same cellular receptors as BHV-1 for viral adsorption and penetration, indicating that the block to BHV-1 infection occurs downstream of entry. However, it is not yet known which mechanisms dictate cellular sensitivity to BHV-1 infection. Additional studies will evaluate the factors determining permissivity to BHV-1 infection, the mechanism of cell death in permissive cell types, and the in vivo tumour killing ability of BHV-1. In the future, these results will contribute to optimizing effectiveness of BHV-1 for use in the oncolytic platform as an ideal cancer therapy. These studies will also provide important insights into the mechanisms that govern virus-host interactions which are crucial for understanding how viruses cause disease and for the development of clinically relevant antiviral treatments.
Breanne Cuddington, Department of Pathology and Molecular Medicine, Institute for Infectious Disease Research, McMaster University, Hamilton, ON, CANADA. In Vitro Cellular and Developmental Biology, 48:S32, 2012
2012 PHILIP R. WHITE AWARD
From Eleanor Vesty:
Eleanor Vesty
The Phillip White Award enabled me to travel to Jena in Germany to work with Dr Thomas Wichard at the Institute for Inorganic and Analytical Chemistry. The institute has well-established protocols and techniques for completion of the life cycle of the alga Ulva mutabilis which is very similar to the U. linza species that is used in my research. I was given in-depth training in these techniques over a period of 3 weeks. There were two main outcomes of the training. One was to observe the various culture methods that Dr Wichard has established to support the algae during different life stages, in order to recreate them in Birmingham. The second was to learn how to induce gametogenesis in a green macroalgal species. I achieved both targets and I am now familiar with the many nuances that determine successful culturing of all stages of the life cycle. I also took samples of U. linza to see if the protocols were transferable and was able to induce gametogenesis in that species as well. This training will allow me to establish a consistent green macroalgal model system in the UK, enabling a suite of other investigative techniques to be carried out. This will be extremely beneficial to my doctoral research and also to the wider Phycological community.
