The Nucleic Acid Fractions of a Strain of Streptococcus faecalis

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Abstract
SUMMARY: The Schmidt & Thannhauser (1945) procedure was applied to the nucleic acid fractions of Streptococcus faecalis. A part of the deoxypentose nucleic acid was differentiated as being insoluble in N-NaOH at 37O, and appeared to be fly bound to polysaccharide material. Base analyses of the nucleic acid fractions are reported, and qualitative determinations of the amino-acid and sugar com- position of the residue described. The Schmidt & Thannhauser (1945) procedure has been widely used to estimate the nucleic acids in biological materials. It consists, essentially, of incubating the tissue with N-NaOH so that the nucleic acids dissolve and the pentose nucleic acid (PNA) is degraded to nucleotides while the deoxypentose nucleic acid (DNA) is still precipitable by acid. The nucleic acid fractions are then resolved by precipitating the DNA, the PNA nucleotides remaining in the supernatant. We have noted, however, in applying this method to a strain of Streptococcus faecalis, that a large proportion of the DNA, in association with much cell material, remains undissolved in the N-NaOH. Thus the DNA portion appears to exist in two easily separable fractions. METHODS Preparation of material. Streptococcus faecalis NCIB 8123 was grown in a medium comprising 0.2 g. yeast extract (Oxoid), 0-5 g. peptone (Oxoid), 0.5 g. glucose, dissolved in 100 ml. 0.05 &I-phosphate buffer and adjusted to pH 7. The organism was grown in 10 1. batches, harvested on a Sharples supercentrifuge, washed twice with water and acetone-dried. Schmidt & Thannhauser procedure. A modification of this technique (David- son, Leslie & Waymouth, 1949) was used to prepare the nucleic acid fractions. The dried cells were extracted twice with ice-cold 10 yo (w/v) trichloroacetic acid (TCA) in 50 ml. centrifuge tubes, then, in succession, with 80 yo (vlv) ethanol, absolute ethanol, chloroform + ethanol (1 : 3) twice at 80°, and finally with ether. The remaining dry powder was incubated overnight with 10 ml. N-NaOH at 37'. The insoluble residue was centrifuged off and washed with water (15 ml.) which was added to the supernatant. To this combined aqueous solution, 5 ml. of 2.5 N-HCI and 6 ml. 30 % (w/v) TCA were added. The DNA was precipitated and spun down by centrifuging for 20 min. at 5000 r.p.m. The precipitate was washed twice with small volumes of 5 yo TCA, the washings being added to the supernatant solution which contained the PNA. This

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