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Northern Analysis: The Basics

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Individually packaged, large volume NorthernMax and NorthernMax-Gly reagents are also available.

Because Northern blots are used extensively to assess mRNA size, RNA markers are necessary for size determination. Ambion offers Millennium Markers, a set of evenly spaced, single-stranded RNA transcripts for accurate determination of mRNA size. The Millennium Marker mixture includes 0.5, 1, 1.5, 2, 2.5, 3, 4, 5, 6, and 9 kilobase transcripts that can be radioactively labeled, probed with our Millennium Marker Probe, or ethidium bromide stained to provide an accurate sizing ladder in gels or on autoradiographs.


Transfer to Solid Support and Immobilization

Once separated by denaturing agarose gel electrophoresis, the RNA is transferred to a positively charged nylon membrane and then immobilized for subsequent hybridization. The best low-tech method for agarose transfer is by a passive, slightly alkaline, downward elution. This procedure, in comparison to upward transfer, is much faster and therefore results in tighter bands and more signal. Alternatively, commercially available active transfer methods (electroblotter, semidry electroblotter, vacuum blotter, pressure blotter, etc.) can be used. For a more detailed
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