Characterization of Deoxyribonucleic Acid from Anacystis nidulans

Abstract

THE chemical nature of deoxyribonucleic acid in blue-green algæ was not known until very recently^1–3. This communication deals with a quantitative study of deoxyribonueleic acid from Anacystis nidulans and its characterization in respect to that from other sources. Anacystis nidulans and Chlorella ellipsoidea were grown in continuous culture chambers as described earlier^4,5. The algal cells were treated with cold 10 per cent trichloracetic acid, 95 per cent alcohol, alcohol/ether (3 : 1) and finally with ether until all the pigments except phycobilins were fully extracted. Deoxyribonucleic acid was isolated by one of the following procedures : (a) with 10 per cent sodium chloride (2 ml./ 10 mgm.) three times by refluxing at 80° C. for 1 hr. each time⁶ ; (b) with 1 N potassium hydroxide at room temperature⁷; (c) with 0.5 M perchloric acid⁸. Purification of deoxyribonueleic acid from protein and polysaccharide was done according to the methods described earlier⁹. Chromatographic analyses were done according to Wyatt¹⁰. Purines and pyrimidines were estimated by maximum extinction technique¹¹. With different procedures of extraction it has been observed that the yield of purified deoxyribonucleic acid varies from 3.7 to 4.7 per cent of the dried cells after ether wash. The fresh weight of the cells was found to be 5 times that of the dried ether-washed cells and hence deoxyribonueleic acid constitutes 0.73–0.94 per cent of the fresh weight of the cells. Nitrogen content varies from 13.9 to 14.3 per cent and that of phosphorus from 6.8 to 8.2 per cent with different procedures of isolation. Deoxyribonucleic acid is found to have an absorption maximum at 257 mµ and minimum at 230 mµ The infra-red spectrum of deoxyribonueleic acid from Anacystis does not show any remarkable deviations from those of other sources. The main peaks are at 3, 6.1, 8.1 and 9.3–9.8µ The percentage of molar ratios of constituent bases, guanine, adenine, cytosine and thymine, in deoxyribonueleic acid of Anacystis was found to be 29.5, 23.3, 24.8, and 22.3 respectively. The ratios of G/C, A/T and Pu/Py were 1.19, 1.04 and 1.12 respectively. The average values only have been recorded here. The deviation of unity in base ratio in this case is very difficult to explain at present, though the presence of single-stranded deoxyribonueleic acid which has recently been isolated from bacteriophage¹² cannot be ruled out. The (A + T)/(G + C) ratio may be used as a species parameter. In the case of Anacystis nidulans this was found to be about 0.82.