Expression in Tobacco Transgenic Plants of a Gene for M2e Peptide of H5NI Avian Influenza Virus in Translational Fusion with Ricin B Subunit

Автор: A.P. Firsov, I.V. Tarasenko, A.S. Pushin, L.A. Shaloiko, L.M. Vinokurov, and S.V. Dolgov

Страница: 55-64

Expression in Tobacco Transgenic Plants of a Gene for M2e Peptide of H5NI Avian Influenza Virus in Translational Fusion with Ricin B Subunit

Biotekhnologiya, 2015, N 2, P. 55-64

UDC 575.113.1:577.218:633.71]:615.371

Section:  “Producers, Biology, Selection, and Gene Engineering”

A.P. Firsov ^*,  I.V. Tarasenko ^*,  A.S. Pushin,  L.A. Shaloiko,  L.M. Vinokurov,  and  S.V. Dolgov

The Institute for Bioorganic Chemistry, Russ. Acad. Sci., Pushchino branch,  142290, Pushchino, Moskovskaya oblast Russia

e-mail:  aleksey_firsov@mail.ru,  starassenko@rambler.ru

The M2e peptide of avian influenza virus H5N1 Curgan 2005 has successfully been expressed in transgenic tobacco plants. The nucleotide sequence encoding the amino-terminal fragment of the protein M2, 30 a.a. in size that included the M2e peptide (M130) was translationally fused within the reading frame with the 3'-terminus of the ricin B subunit (RTB) sequence in vector pBI121. Tobacco plants were successfully transformed with the obtained construct. The RTB was detected in 4 lines of the transgenic plants from the 8 studied using asialofetuin-immobilized ELISA. The presence of the M2e peptide in the fused protein RTB-M130 was confirmed in two lines of the transgenic plants; in 2 other lines M2e was not detected regardless of the presence of RTB. The quantitative asialofetuin-immobilized ELISA permitted to show that the accumulation of the fused protein RTB-M130 in those lines was equal to 3.3 and 2.4 µg per 1 g of leaf fresh weight (0.02% and 0.01% of total soluble protein, respectively). The lines in which the expression of the RTB gene was only detected were characterized by the RTB accumulation of 2.1 and 2.4 µg per 1 g of fresh weight (0.01 and 0.02% of the total soluble protein, respectively). Western blot analysis showed that the RTB-M130 protein was detected in the transgenic tobacco lines as a dimer without significant signs of degradation. The obtained results will be further used for the design of a plant-based edible vaccine against avian influenza.

Key words:  avian influenza,  edible vaccines,  molecular farming,  peptide M2e,  ricin B subunit.

25.06.2015, 1470 просмотров.