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Ribosome profiling of mouse hepatitis virus (MHV)-infected and uninfected 17Cl1 cells

ABSTRACT: Ribosome profiling of mouse hepatitis virus (MHV)-infected and uninfected 17Cl1 cells

PROVIDER: PRJEB23773 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR2213251.fastq.gz	Fastqsanger.gz
	ERR2213252.fastq.gz	Fastqsanger.gz
	ERR2213253.fastq.gz	Fastqsanger.gz
	ERR2213254.fastq.gz	Fastqsanger.gz
	ERR2213255.fastq.gz	Fastqsanger.gz

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Ribosome profiling of mouse hepatitis virus (MHV)-infected and uninfected 17Cl1 cells

Project description:Ribosome profiling (RiboSeq) analysis of murine 17 clone 1 (17Cl-1) cells with and without MHV infection. We sought to assess the impact of differing library preparation methods by using three separate approaches: flash freezing, 1X cycloheximide pretreatment, and 100X cycloheximide pretreatment.

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RiboSeq of murine 17Cl-1 cells infected at MOI 10 with MHV-A59, compared to mock-infected controls, harvested at 5 and 8 hpi, plus a control tunicamycin-treated sample.

Project description:Murine 17Cl-1 cells were infected with the model Betacoronavirus mouse hepatitis virus (MHV) strain A59 and subjected to ribosome profiling to observe changes in the host translatome in infected cells compared to mock-infected cells. Samples were harvested at 5 and 8 hpi by flash-freezing, without cycloheximide pre-treatment. RNase I treatment was carried out, following which ribosomes and enclosed RNA were isolated by centrifugation through a sucrose cushion. RNA was extracted, ribosomal RNA was removed using Illumina's RiboZero kit, and remaining RNA was gel purified to select fragments 25-35nt long. Fragments were cloned into adapters using the TruSeq small RNA adapter kit and sequenced on Illumina NextSeq, then differential translation efficiency analysis was carried out on the data deposited data. One uninfected sample was treated with 2ug/mL tunicamycin for 6 hours before harvesting as a positive control for unfolded protein response activation.

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