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URL: https://dx.doi.org/10.17504/protocols.io.bd9ti96n

Authors: Ariel Rabines, Rob Lampe, Andrew Allen

Group: A.E. Allen Lab

Summary: A mostly automated protocol for extraction of total RNA from seawater filtered onto a Sterivex filter (Cat. No. SVGP0150). Reagents come from the Macherey-Nagel NucleoMag RNA kit (Cat. No. 744350). Automated liquid handling is performed on an eppendorf EpMotion 5075t with multi-channel pipettes. This protocol assumes that the Sterivex is sealed with tube sealant on the male end and a male-luer lock plug on the female end. Using this protocol, we routinely extract ~80 samples at a time.The protocol has been modified to increase to an 800 µL starting volume. This normally results in less Lysis Buffer MR1, Binding Buffer MR2, and Reducing Agent TCEP supplied than is needed. Extra reagents can and should be purchased separately. The kit manual can be found here: https://www.mn-net.com/media/pdf/23/c2/23/Instruction-NucleoMag-RNA.pdfAdditional notes from the manufacturer on using the kit with the epMotion can be found here: https://www.mn-net.com/media/pdf/bd/7e/53/AN-NucleoMag-RNA-epMotion5075.pdf . Note that we have several additional modifications to their protocol.The epMotion program is attached here.

Affiliations: Scripps Institution of Oceanography, Scripps Institution of Oceanography, Scripps Institution of Oceanography

Version: 1

Publication Date: 2020

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