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Cold Spring Harb Protoc. 2017 Apr 3;2017(4):pdb.prot091629. doi: 10.1101/pdb.prot091629.

Preparation of Total RNA from Fission Yeast.

Author information

1
Department of Genetics, Evolution & Environment, and UCL Cancer Institute, University College London, London WC1E 6BT, United Kingdom; j.bahler@ucl.ac.uk.
2
Center for RNA Molecular Biology and Department of Biochemistry, Case Western Reserve University, Cleveland, Ohio 44106-4906.

Abstract

Treatment with hot phenol breaks open fission yeast cells and begins to strip away bound proteins from RNA. Deproteinization is completed by multiple extractions with chloroform/isoamyl alcohol and separation of the aqueous and organic phases using MaXtract gel, an inert material that acts as a physical barrier between the phases. The final step is concentration of the RNA by ethanol precipitation. The protocol can be used to prepare RNA from several cultures grown in parallel, but it is important not to process too many samples at once because delays can be detrimental to RNA quality. A reasonable number of samples to process at once would be three to four for microarray or RNA sequencing analyses and six for preliminary investigations of mutants implicated in RNA metabolism.

PMID:
28373489
DOI:
10.1101/pdb.prot091629
[Indexed for MEDLINE]

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