High-Effective Production of Biologically Active Secreted Human Granulocyte Macrophage Colony-Stimulating Factor by Recombinant Vaccinia Virus

Автор: A.A. Grazhdantseva, G.F. Sivolobova, A.V. Tkacheva, I.P. Gileva, E.V. Kuligina, V.A. Rikhter, and G.V. Kochneva

Страница: 13-21

High-Effective Production of Biologically Active Secreted Human Granulocyte Macrophage Colony-Stimulating Factor by Recombinant Vaccinia Virus

Biotekhnologiya, 2015, N 5, P. 13-21

UDC 577.21:578.28: 615.371

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

A.A. Grazhdantseva ^{1, 2},  G.F. Sivolobova ^{1, 2},  A.V. Tkacheva ^{1, 2},  I.P. Gileva ¹,  E.V. Kuligina ²,  V.A. Rikhter ²,  and  G.V. Kochneva ^{1, 2, *}

¹  The State Research Center for Virology and Biotechnology Vector,  Novosibirskaya oblast, Kol'tsovo, 630559 Russia

²  The Institute for Chemical Biology and Fundamental Medicine, Russ. Acad. Sci., Siberian branch,  Novosibirsk, 630090 Russia

e-mail:  g.v.kochneva@yandex.ru

A recombinant vaccinia virus strain VV-GMCSF-S1/3 that contains an insertion of a full-length DNA copy of messenger RNA of human granulocyte-macrophage colony-stimulating factor (GM-CSF) in the structural part of the viral thymidine kinase gene has been obtained. The expression of the GM-CSF gene as a part of the recombinant virus is under the control of the native vaccinia virus promoter р7.5K and results in the production of the mature form of the secreted protein with a molecular mass of 32 kDa. The biological activity of the GM-CSF was evaluated from the stimulation of the cytokine-dependent human erythroleukemia TF-1 cell proliferation. The level of the secretion of the biologically active human GM-CSF in the recombinant vaccinia virus / mammalian cells system was 1—40 μg/ml of culture medium. The recombinant strain of VV-GMCSF-S1/3 can be used as a producer of the glycosylated mature form of human GM-CSF, as well as a vector for the construction of oncolytic viruses and multivalent vaccine preparations.

Key words:  cell proliferation assay XTT,  human erythroleukemia TF-1 cells,  GM-CSF,  recombinant vaccinia virus,  strain L-IVP.

20.05.2016, 1384 просмотра.