[INCLUSION OF SITE-SPECIFIC MUTATIONS INTO CONSERVATIVE SEG- MENTS OF PA-GENE RESULTS IN ATTENUATION OF VIRULENT INFLUENZA VIRUS STRAIN A/WSN/33].

Autor:	Rtischev AA, Mintaev RR, Kost VY, Koptyaeva LB, Akopova II, Lisovskaya KV, Markushin SG
Jazyk:	English; Russian
Zdroj:	Zhurnal mikrobiologii, epidemiologii i immunobiologii [Zh Mikrobiol Epidemiol Immunobiol] 2017 Mar (2), pp. 45-53.
Abstrakt:	Aim: Study the possibility of obtaining attenuated variants of influenza virus by including specially selected site-specific mutations into a conservative sequence of PA-gene (terminal segment of COOH-domain of the PA-gene) of a virulent strain. Materials and Methods: A/ WSN/33 - a virulent strain of influenza virus was used in the study. Inclusion of site-specif- ic mutations into PA-gene of the A/WSN/33 virulent strain was carried out using a two-step mutation PCR. Cloning was carried out using GoldenGate reaction. 8-plasmid transfection system based on pHW2000 vector was used. Transformation was carried out in rubidium competent bacterial cells of DH5(α strain. Transfection was done using Lipofectamine LTX (Invitrogen) reagentin a 293T and MDCK cells' co-culture. Results: Transfectants with F658A substitution in the COOH-domain of the PA-gene were shown to acquire ts-phenotype and sharply reduce the ability to reproduce in mice lungs. Introduction of F658A substitution into COOH-domain of the PA-gene in combination with introduction of ts-mutations from ca influenzavirus strains into the genome ofthe virulent strain resulted in obtaining transfectants that have phenotypic characteristics typical for live influenza vaccine candidates. Conclusion: The ability to obtain attenuated variants of influenza viruses by introducing spe- cially selected site-specific mutations into conservative sequence of the PA-gene is shown.
Databáze:	MEDLINE
Externí odkaz:	Zobrazit plný text záznamu