Document Detail

Biosynthetic preparation of 13C/15N-labeled rNTPs for high-resolution NMR studies of RNAs.
MedLine Citation:
PMID:  23065565     Owner:  NLM     Status:  MEDLINE    
High-resolution investigations of the structure and dynamics of RNA molecules by nuclear magnetic resonance (NMR) methodologies require the production of (13)C/(15)N-isotopically labeled samples. A common strategy entails the preparation of (13)C/(15)N-enriched ribonucleoside 5'-triphosphates (rNTPs) to be incorporated into RNA oligomers by in vitro transcription. Here, we describe the methods to obtain isotopically labeled rNTP in a uniform or selective fashion from bacterial cultures, using common and versatile E. coli strains. This chapter also covers procedures for extraction and digestion of the total RNA from bacterial cells, purification of the ribonucleoside 5'-monophosphates and their enzymatic phosphorylation to rNTPs.
Luigi Martino; Maria R Conte
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Methods in molecular biology (Clifton, N.J.)     Volume:  941     ISSN:  1940-6029     ISO Abbreviation:  Methods Mol. Biol.     Publication Date:  2012  
Date Detail:
Created Date:  2012-10-15     Completed Date:  2013-03-18     Revised Date:  2014-02-20    
Medline Journal Info:
Nlm Unique ID:  9214969     Medline TA:  Methods Mol Biol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  227-45     Citation Subset:  IM    
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MeSH Terms
Acetates / pharmacology
Carbon Isotopes / chemistry
Cell Proliferation
Culture Media / chemistry
Escherichia coli / cytology,  drug effects,  genetics,  metabolism
Formates / pharmacology
Glucose / pharmacology
Glycerol / pharmacology
Isotope Labeling
Magnetic Resonance Spectroscopy
Nitrogen Isotopes / chemistry
RNA / biosynthesis*,  chemistry*
Ribonucleotides / biosynthesis*,  chemistry*
Substrate Specificity
Transcription, Genetic
Grant Support
MC_U117533887//Medical Research Council; //Wellcome Trust
Reg. No./Substance:
0/Acetates; 0/Carbon Isotopes; 0/Culture Media; 0/Formates; 0/Nitrogen Isotopes; 0/Ribonucleotides; 63231-63-0/RNA; 64-18-6/formic acid; IY9XDZ35W2/Glucose; PDC6A3C0OX/Glycerol

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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