In situ dot blots: quantitation of mRNA in intact cells.
A rapid, simple and reproducible dot blot method is described for quantitating the amounts of specific messages in small numbers of intact cells. The method has been used to accurately determine the number of histone H4 mRNA molecules in growing (approximately 40,000) and in starved (approximately 1600) Tetrahymena thermophila, and to measure the amount of message contributed by an E. coli plasmid containing part of the S10 ribosomal operon. Use of the method is illustrated to optimize in situ hybridization protocols and to measure mRNA amounts in cell lysates. Preliminary studies also indicate that the method can be used to detect mRNA in intact yeast cells.
D019286:Endopeptidase K; D010450:Endopeptidases / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics; D005976:Glutaral / Q000494:pharmacology; D006657:Histones / Q000235:genetics; D007700:Kinetics; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000032:analysis*; D013768:Tetrahymena / Q000235:genetics; D015003:Yeasts / Q000235:genetics
D006657:Histones; D012333:RNA, Messenger; D010450:Endopeptidases; D019286:Endopeptidase K; D005976:Glutaral