NUCLEOTIDES I3I 



and chromatography, both on paper [11] and on ion ex- 

 change resins [18], have provided more convenient and pre- 

 cise techniques for the separation, preparation [19] and 

 identification of nucleotides, nucleosides and the bases. 

 When a paper chromatogram is exposed to light of wave- 

 length 260 m^w, the areas occupied by purines, pyrimidines, 

 nucleosides or nucleotides appear as dark spots on a light 

 blue fluorescent background. A permanent photographic 

 record can be obtained by placing the chromatogram over 

 reflex copying paper [37], and the areas in the chromato- 

 gram containing compounds absorbing ultraviolet light will 

 appear as white spots on a dark background in the developed 

 photograph (Plate II). The appropriate areas of the chro- 

 matogram are then cut out, the compounds eluted and 

 estimated spectrophotometrically. 



Microbial nucleic acids 



Micro-organisms, particularly bacteria, are richer in 

 nucleic acid than most of the cells of other organisms, and 

 Belozersky has calculated that 15-30% of the dry weight of 

 bacteria is nucleic acid and 50-80% is nucleoprotein [4]. 

 Prior to isolating a nucleic acid, soluble nucleotides and 

 phospholipoids are first removed by successively extracting 

 the cells with cold TCA and a fat solvent. The residue is 

 then treated with dilute solutions of an alkali, e.g. 0-2% 

 NaOH or NagCOg in order to extract the nucleic acids, the 

 details of the procedure varying according to the nucleic 

 acid required. The isolation of microbial nucleoproteins, 

 especially in an undegraded or 'native' state, presents many 

 difficulties and, apart from low yields, it is doubtful if any 

 of the present techniques are ideal. The most favoured 

 method is to extract the cells with neutral solutions of NaCl 

 at a concentration depending on the type of nucleic acid it 

 is desired to isolate: even so, the deoxypentose nucleopro- 

 teins adhere strongly to the cell structure and are only re- 

 moved with difficulty [4]. When an aqueous solution of a 

 nucleoprotein is shaken with chloroform, the protein be- 

 comes denatured and collects at the interface, whilst the 

 liberated nucleic acid remains in the water phase. The 



