The Northern blotting technique involves transfer of RNA species separated in electrophoretic gels to membrane filters for detection of specific base sequences by complementary probes. Northern blots are used to identify and quantitate specific RNA sequences.

1. Inappropriate membrane was used.
2. Immobilization of nucleic acids to membrane was not efficient.
3. Probe concentration was too low.
4. Hybridization and washing conditions were too stringent

This is the most common problem with Northern blots.

1. Concentration of probe was too high.
2. Hybridization and washing conditions were not appropriate.
3. Blocking was inadequate.
4. Washing was inadequate.
5. Target concentration was too high.
6. There was non-specific binding of detection reagents to the membrane

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