Northern blot detection of endogenous small RNAs (approximately14 nt) in bacterial total RNA extracts.
Here we describe a northern blot procedure that allows the detection of endogenous RNAs as small as approximately 14 nt in total RNA extracts from bacteria. RNAs that small and as part of total bacterial RNA extracts usually escape detection by northern blotting. The approach combines LNA probes 5'-digoxigenin-endlabeled for non-radioactive probe detection with 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide-mediated chemical crosslinking of RNAs to nylon membranes, and necessitates the use of native PAGE either with the TBE or MOPS buffer system.
Beckmann
Benedikt M
BM
Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Marburg, Germany.
Grünweller
Arnold
A
Weber
Michael H W
MH
Hartmann
Roland K
RK
D001412:Bacillus subtilis / Q000235:genetics; D001483:Base Sequence; D015152:Blotting, Northern / Q000379:methods*; D002021:Buffers; D003432:Cross-Linking Reagents; D004926:Escherichia coli / Q000235:genetics; D005022:Ethyldimethylaminopropyl Carbodiimide / Q000737:chemistry; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009691:Nucleic Acid Denaturation; D009843:Oligoribonucleotides / Q000737:chemistry; D012329:RNA, Bacterial / Q000032:analysis* / Q000737:chemistry; D022661:RNA, Untranslated / Q000032:analysis* / Q000737:chemistry
D002021:Buffers; D003432:Cross-Linking Reagents; D009843:Oligoribonucleotides; D012329:RNA, Bacterial; D022661:RNA, Untranslated; D005022:Ethyldimethylaminopropyl Carbodiimide