A simple method to enrich mRNA from total prokaryotic RNA.
Isolation of prokaryotic mRNA by the poly(dT) method has been difficult, primarily due to the great instability of the poly(A) sequence in its mRNA. We developed a simple method to remove rRNA from total RNA of Staphylococcus aureus by cloning a PCR-amplified S. aureus rRNA gene fragment into a plasmid, and then synthesizing biotin-labeled antisense rRNA to subtract rRNA. By using this method, S. aureus rRNA is significantly reduced and mRNA is enriched. This method may be used to prepare prokaryotic mRNA for many molecular biology applications.
Dept. of Veterinary Science, Pennsylvannia State University, University Park 16802, USA.
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D003001:Cloning, Molecular; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods*; D011387:Prokaryotic Cells / Q000737:chemistry*; D016372:RNA, Antisense; D012333:RNA, Messenger / Q000302:isolation & purification*; D012335:RNA, Ribosomal / Q000235:genetics / Q000302:isolation & purification; D013211:Staphylococcus aureus / Q000235:genetics
D016372:RNA, Antisense; D012333:RNA, Messenger; D012335:RNA, Ribosomal