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Separation of differentially isotope-labeled bacterial RNA by isopycnic density gradient centrifugation is a critical step in RNA-based stable isotope probing analyses, which help to link the structure and function of complex microbial communities. Using isotope-labeled Escherichia coli RNA, we showed that an 8 mL near-vertical rotor performed better than a 2 mL fixed-angle rotor, thereby corroborating current recommendations. Neither increased concentrations of formamide nor urea in the medium improved the separation results using the fixed-angle rotor.

Effect of rotor type on the separation of isotope-labeled and unlabeled Escherichia coli RNA by isopycnic density ultracentrifugation