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224 Production of HIV-1 Vaccine Components in Plants
A. V. Karasev*, B. V. Kim, K. J. Shon, and H. Koprowski
Thomas Jefferson Univ., Doylestown, PA, USA
Background: Plants represent safe and inexpensive delivery vehicles for components of a prospective HIV-1 vaccine, in particular in the poorest and least developed countries in the world. For instance, success of any future HIV-1 vaccine in combating AIDS in Africa will be dependent to a large extent on economics of its delivery. Our research is focused on production of HIV-1 vaccine components in plants with the emphasis on edible plants, like spinach. The tat protein has recently become a focus of vaccine research as a potential target for a broad, subtype-nonspecific HIV-1 vaccine which may be suitable for harsh conditions in Africa.
Methods: The full-length tat gene of the MN strain of HIV-1 was assembled using synthetic primers, cloned into a tobacco mosaic virus (TMV)-based vector, expressed the tat protein in plants, and developed plant-based production of the tat protein. Several constructs were designed expressing either tat protein alone or as fusions with different stabilizing tags or plant virus capsid proteins used as carriers.
Results: The chimeric TMV-derived constructs were shown to replicate successfully in inoculated leaves of Nicotiana benthamiana and spinach. The tat protein gene, when expressed alone caused no or very mild symptoms in N. benthamiana, and no symptoms in spinach. When fused to plant virus capsid proteins, tat protein gene-containing constructs caused stunting of N. benthamiana plants, leaf curling and mosaic; whereas the same constructs caused only mild yellowing reaction in spinach leaves. The yield of tat protein expressed both in N. benthamiana and spinach plants was estimated to exceed 300 mug of extractable protein per 1 g of the leaf tissue. The plant-expressed tat protein fully retained immunological reactivity against tat-specific monoclonal antibodies.
Conclusions: The availability of this plant-based, inexpensive, and easily scalable production system, which can produce fully immunogenic tat protein is an important step toward HIV-1 vaccine.
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