Dataset Information

Lentviral-mediated RNAi to inhibit target gene expression of the porcine integrin ?v subunit, the FMDV receptor, and against FMDV infection in PK-15 cells.

ABSTRACT: BACKGROUND: shRNA targeting the integrin ?v subunit, which is the foot-and-mouth disease virus (FMDV) receptor, plays a key role in virus attachment to susceptible cells. We constructed a RNAi lentiviral vector, i?v pLenti6/BLOCK -iT™, which expressed siRNA targeting the FMDV receptor, the porcine integrin ?v subunit, on PK-15 cells. We also produced a lentiviral stock, established an i?v-PK-15 cell line, evaluated the gene silencing efficiency of mRNA using real-time qRT-PCR, integrand ?v expression by indirect immunofluorescence assay (IIF) and cell enzyme linked immunosorbent assays (cell ELISA), and investigated the in vivo inhibitory effect of shRNA on FMDV replication in PK-15 cells. RESULTS: Our results indicated successful establishment of the i?v U6 RNAi entry vector and the i?v pLenti6/BLOCK -iT expression vector. The functional titer of obtained virus was 1.0 × 10(6) TU/mL. To compare with the control and mock group, the i?v-PK-15 group ?v mRNA expression rate in group was reduced by 89.5%, whilst IIF and cell ELISA clearly indicated suppression in the experimental group. Thus, i?v-PK-15 cells could reduce virus growth by more than three-fold and there was a > 99% reduction in virus titer when cells were challenged with 10(2) TCID(50) of FMDV. CONCLUSIONS: I?v-PK-15 cells were demonstrated as a cell model for anti-FMDV potency testing, and this study suggests that shRNA could be a viable therapeutic approach for controlling the severity of FMD infection and spread.

SUBMITTER: Luo J

PROVIDER: S-EPMC3183033 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Lentviral-mediated RNAi to inhibit target gene expression of the porcine integrin αv subunit, the FMDV receptor, and against FMDV infection in PK-15 cells.

Luo Jihuai J Du Junzheng J Gao Shandian S Zhang Guofeng G Sun Jingjing J Cong Guozheng G Shao Junjun J Lin Tong T Chang Huiyun H

Virology journal 20110907

<h4>Background</h4>shRNA targeting the integrin αv subunit, which is the foot-and-mouth disease virus (FMDV) receptor, plays a key role in virus attachment to susceptible cells. We constructed a RNAi lentiviral vector, iαv pLenti6/BLOCK -iT™, which expressed siRNA targeting the FMDV receptor, the porcine integrin αv subunit, on PK-15 cells. We also produced a lentiviral stock, established an iαv-PK-15 cell line, evaluated the gene silencing efficiency of mRNA using real-time qRT-PCR, integrand α ...[more]

PMID: 21899738

Similar Datasets

Project description:Porcine deltacoronavirus (PDCoV) is a newly emerged swine enteropathogenic coronavirus affecting pigs of all ages and causing diarrhea problems. Research findings indicate that PDCoV has evolved strategies to escape innate immune response in host cells, but mechanism of PDCoV in innate immune modulation is not well understood. In this study, we report our findings on identifying the alterations of host cell innate immune response affected by PDCoV infection and exploring the gene expression profiles of PK-15 cells at 0, 24, and 36 h PDCoV post infection by RNA sequencing. A total of 3,762 and 560 differentially expressed genes (DEGs) were screened by comparison of uninfected PK-15 cells and infected PK-15 cells at 24 h post infection (hpi) (INF_24h versus NC), and also comparison of infected PK-15 cells between 24 and 36 hpi (INF_36h versus INF_24h), which included 156 and 23 porcine innate immune-related genes in the DEGs of INF_24h versus NC and INF_36h versus INF_24h, respectively. Gene Ontology function classification and Kyoto Encyclopedia of Genes and Genomes signaling pathway enrichment analysis were performed based on the DEGs that exhibited the same expression tendencies with most of the innate immune-associated genes among these PK-15 cell samples described above. The enrichment results indicated that extensive gene functions and signaling pathways including innate immune-associated functions and pathways were affected by PDCoV infection. Particularly, 4 of 5 innate immune signaling pathways, which were primarily affected by PDCoV, played important roles in I-IFN's antiviral function in innate immune response. Additionally, 16 of the host cell endogenous miRNAs were predicted as potential contributors to the modulation of innate immune response affected by PDCoV. Our research findings indicated that the innate immune-associated genes and signaling pathways in PK-15 cells could be modified by the infection of PDCoV, which provides a fundamental foundation for further studies to better understand the mechanism of PDCoV infections, so as to effectively control and prevent PDCoV-induced swine diarrheal disease outbreaks.

Dataset Information

Lentviral-mediated RNAi to inhibit target gene expression of the porcine integrin ?v subunit, the FMDV receptor, and against FMDV infection in PK-15 cells.

Publications

Lentviral-mediated RNAi to inhibit target gene expression of the porcine integrin αv subunit, the FMDV receptor, and against FMDV infection in PK-15 cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets