Abstract
Publisher Summary This chapter discusses 16 S ribosomal RNA cataloging. Phylogenetic studies on cyanobacteria and chloroplasts using the 16 S ribosomal RNA (rRNA) cataloging approach as a tool have been done by two different methods: the traditional one which is generally not used anymore and the advanced technique which is presented in detail in chapter. rRNA can be isolated either from ribosomal subunits or from crude extracts. Experience with more than 450 prokaryotic species has shown that the following steps will lead in most cases to satisfactory amounts and purity of 16 S rRNA. The original methods of sequence determination have been improved to facilitate sequencing of the more than 70 oligonucleotides, varying in length from 6 to 20 nucleotides, making up a 16 S rRNA catalog. These changes include the replacement of the “Homomixes” by ammonium formate gradient chromatography, the use of alkaline hydrolysis rather than enzymatic cleavage for the generation of subfragments, and the introduction of additional steps to improve analysis of those oligonucleotides whose sequence could not be determined unambiguously after the secondary analysis.
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