Comparison of translational enhancers in the course of in vitro synthesis of sheep pox viral coat proteins A27L and L1R
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Keywords:
enhancers of mRNA translation, plant cell-free system, synthesis in vitro of coat proteins A27L and L1R, sheep pox virus.Abstract
In this work the abilities of plant viral and artificial enhancers of mRNA translation to increase synthesis of sheep pox viral coat proteins A27L and L1R in wheat germ cell-free system is studied and compared. For such a comparison 5' untranslated regions (UTRs) of genomic (g)RNAs of potato virus Y (PVY), tobacco etch virus (TEV), gRNA-4 of alfalfa mosaic virus (AMV), artificial enhancer 5xARC1, as well as 3' UTR of a tobacco mosaic virus (TMV) gRNA were used. The assessment of translation enhancement was made by measurement of activity of newly synthesized reporter protein β-glucuronidase (GUS) that hydrolyzes the fluorogenic substrate, as well as with the help of immunoblotting of proteins A27L and L1R. It is established that examined 5'-UTRs can considerably raise synthesis of proteins in comparison with arbitrary leader sequence (pl), which does not possess translational enhancer property.References
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2 Hellen Ch.U.T., Sarnow P. Internal ribosome entry sites in eukaryotic mRNA molecules // Genes Dev. – 2001. –
V.15. – P.1593-1612.
3 Орлова Е.С. Совершенствование методов диагностики оспы овец и оспы коз. – Диссертация на соискание
ученой степени кандидата биологических наук. –Владимир, 2007. – 6 с.
4 Mangana-Vougiouka, O. Sheep poxvirus identification from clinical specimens by PCR, cell culture,
immunofluorescence and agar gel immunoprecipitation assay // Molecular and Cellular Probes. – 2000. – V.14. -No5. –P.305-310.
5 Gallie D.R., Feder J.N., Schimke R.T., Walbot V. Post-transcriptional regulation in higher eukaryotes: The role of the
reporter gene in controlling expression // Mol.Gen.Genet. – 1991. –V.228. – P. 258-264.
6 Gurevich. V., Pokrovskaya I.D., Obukhova T.A. Preparative in vitro mRNA synthesis using SP6 and T7 RNA
polymerases // Anal.Biochem. – 1991. –V.195. – P. 207-213.
7 Akbergenov R.Zh., Zhanybekova S.Sh., Kryldakov R.V., Zhigailov A.V., Polimbetova N.S., Hohn T., Iskakov B.K.
ARC-1, a sequence element complementary to an internal 18S rRNA segment, enhances translation efficiency in plants
when present in the leader or intercistronic region of mRNAs // Nucleic Acids Research. – 2004. – V.32. – No.1. – P.239-247.
8 Johnston F.B., Stern H. Mass isolation of viable wheat embryo // Nature. – 1957. – No.179. – P. 160-161.
9 Maniatis T., Fritsch E.F., Sambrook J. Molecular cloning (a laboratory manual). -New York: Cold Spring Harbor
Lab.Press, 1989, V.3. – 545 р.
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Beisenov, D. K., Stanbekova, G. E., Zhigailov, A. V., & Iskakov, B. K. (2015). Comparison of translational enhancers in the course of in vitro synthesis of sheep pox viral coat proteins A27L and L1R. Experimental Biology, 59(3/1), 49–52. Retrieved from https://bb.kaznu.kz/index.php/biology/article/view/643
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