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Int J Syst Bacteriol 39 (1989), 159-167; DOI 10.1099/00207713-39-2-159
© 1989 Society for General Microbiology
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Precise Measurement of the G+C Content of Deoxyribonucleic Acid by High-Performance Liquid Chromatography

MOSTAFA MESBAH{dagger}, USHA PREMACHANDRAN and WILLIAM B. WHITMAN*

Department of Microbiology, University of Georgia, Athens, Georgia 30602

* Corresponding author.

ABSTRACT

High-performance liquid chromatography is a promising alternative for determining the G+C content of bacterial deoxyribonucleic acid (DNA). The method which we evaluated involves enzymatic degradation of the DNA to nucleosides by P1 nuclease and bovine intestinal mucosa alkaline phosphatase, separation of the nucleosides by high-performance liquid chromatography, and calculation of the G+C content from the apparent ratios of deoxyguanosine and thymidine. Because the nucleosides are released from the DNA at different rates, incomplete degradation produces large errors in the apparent G+C content. For partially purified DNA, salts are a major source of interference in degradation. However, when the salts are carefully removed, the preparation and degradation of DNA contribute little error to the determination of G+C content. This method also requires careful selection of the chromatographic conditions to ensure separation of the major nucleosides from the nucleosides of modified bases and precise control of the flow rates. Both of these conditions are achievable with standard equipment and C18 reversed-phase columns. Then the method is precise, and the relative standard deviations of replicate measurements are close to 0.1%. It is also rapid, and a single measurement requires about 15 min. It requires small amounts of sample, and the G+C content can be determined from DNA isolated from a single bacterial colony. It is not affected by contamination with ribonucleic acid. Because this method yields a direct measurement, it may also be more accurate than indirect methods, such as the buoyant density and thermal denaturation methods. In addition, for highly purified DNA, the extent of modification can be determined.


{dagger} Present address: Department of Pharmacognosy, Faculty of Pharmacy, University of Assiut, Assiut, Egypt.




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Int. J. Syst. Evol. Microbiol.Home page
H.-Y. Weon, B.-Y. Kim, C.-M. Lee, S.-B. Hong, Y.-A. Jeon, B.-S. Koo, and S.-W. Kwon
Solitalea koreensis gen. nov., sp. nov. and the reclassification of [Flexibacter] canadensis as Solitalea canadensis comb. nov.
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 1969 - 1975.
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Int. J. Syst. Evol. Microbiol.Home page
S.-K. Tang, Y. Wang, Y. Chen, K. Lou, L.-L. Cao, L.-H. Xu, and W.-J. Li
Zhihengliuella alba sp. nov., and emended description of the genus Zhihengliuella
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 2025 - 2032.
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Int. J. Syst. Evol. Microbiol.Home page
M. Labrenz, P. A. Lawson, B. J. Tindall, and P. Hirsch
Roseibaca ekhonensis gen. nov., sp. nov., an alkalitolerant and aerobic bacteriochlorophyll a-producing alphaproteobacterium from hypersaline Ekho Lake
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 1935 - 1940.
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Int. J. Syst. Evol. Microbiol.Home page
I. Vaz-Moreira, A. R. Lopes, C. Faria, C. Sproer, P. Schumann, O. C. Nunes, and C. M. Manaia
Microbacterium invictum sp. nov., isolated from homemade compost
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 2036 - 2041.
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Int. J. Syst. Evol. Microbiol.Home page
J. Killer, J. Kopecny, J. Mrazek, V. Rada, O. Benada, I. Koppova, J. Havlik, and J. Straka
Bifidobacterium bombi sp. nov., from the bumblebee digestive tract
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 2020 - 2024.
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Int. J. Syst. Evol. Microbiol.Home page
S.-H. Yoo, H.-Y. Weon, R. Anandham, B.-Y. Kim, S.-B. Hong, Y.-A. Jeon, B.-S. Koo, and S.-W. Kwon
Dokdonella soli sp. nov., a gammaproteobacterium isolated from soil
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 1965 - 1968.
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Int. J. Syst. Evol. Microbiol.Home page
Y. Wang, S.-K. Tang, K. Lou, P.-H. Mao, X. Jin, C.-L. Jiang, L.-H. Xu, and W.-J. Li
Paracoccus saliphilus sp. nov., a halophilic bacterium isolated from a saline soil
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 1924 - 1928.
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Int. J. Syst. Evol. Microbiol.Home page
Y.-G. Chen, X.-L. Cui, Y.-X. Wang, Y.-Q. Zhang, S.-K. Tang, W.-J. Li, Z.-X. Liu, M.-L. Wen, and Q. Peng
Virgibacillus sediminis sp. nov., a moderately halophilic bacterium isolated from a salt lake in China
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 2058 - 2063.
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Int. J. Syst. Evol. Microbiol.Home page
A. B. Arun, W.-M. Chen, W.-A. Lai, J.-H. Chou, F.-T. Shen, P. D. Rekha, and C.-C. Young
Lutaonella thermophila gen. nov., sp. nov., a moderately thermophilic member of the family Flavobacteriaceae isolated from a coastal hot spring
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 2069 - 2073.
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Int. J. Syst. Evol. Microbiol.Home page
S. T. Khan, S. Harayama, T. Tamura, K. Ando, M. Takagi, and S.-y. Kazuo
Paraoerskovia marina gen. nov., sp. nov., an actinobacterium isolated from marine sediment
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 2094 - 2098.
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Int. J. Syst. Evol. Microbiol.Home page
N. M. Mesbah and J. Wiegel
Natronovirga wadinatrunensis gen. nov., sp. nov. and Natranaerobius trueperi sp. nov., halophilic, alkalithermophilic micro-organisms from soda lakes of the Wadi An Natrun, Egypt
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 2042 - 2048.
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Int. J. Syst. Evol. Microbiol.Home page
P. Anil Kumar, T. N. R. Srinivas, S. Takaichi, T. Maoka, Ch. Sasikala, and Ch. V. Ramana
Phaeospirillum chandramohanii sp. nov., a phototrophic alphaproteobacterium with carotenoid glycosides
Int J Syst Evol Microbiol, August 1, 2009; 59(8): 2089 - 2093.
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Int. J. Syst. Evol. Microbiol.Home page
Y.-X. Wang, Z.-G. Wang, J.-H. Liu, Y.-G. Chen, X.-X. Zhang, M.-L. Wen, L.-H. Xu, Q. Peng, and X.-L. Cui
Sediminimonas qiaohouensis gen. nov., sp. nov., a member of the Roseobacter clade in the order Rhodobacterales
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1561 - 1567.
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Int. J. Syst. Evol. Microbiol.Home page
C. Y. Hwang, G. D. Bae, W. Yih, and B. C. Cho
Marivita cryptomonadis gen. nov., sp. nov. and Marivita litorea sp. nov., of the family Rhodobacteraceae, isolated from marine habitats
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1568 - 1575.
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Int. J. Syst. Evol. Microbiol.Home page
R. H. Gregersen, C. Neubauer, H. Christensen, A. M. Bojesen, M. Hess, and M. Bisgaard
Comparative studies on [Pasteurella] testudinis and [P.] testudinis-like bacteria and proposal of Chelonobacter oris gen. nov., sp. nov. as a new member of the family Pasteurellaceae
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1583 - 1588.
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Int. J. Syst. Evol. Microbiol.Home page
R. Beaz Hidalgo, I. Cleenwerck, S. Balboa, S. Prado, P. De Vos, and J. L. Romalde
Vibrio breoganii sp. nov., a non-motile, alginolytic, marine bacterium within the Vibrio halioticoli clade
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1589 - 1594.
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Int. J. Syst. Evol. Microbiol.Home page
Y.-R. Cao, Y. Jiang, L.-H. Xu, and C.-L. Jiang
Sphaerisporangium flaviroseum sp. nov. and Sphaerisporangium album sp. nov., isolated from forest soil in China
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1679 - 1684.
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Int. J. Syst. Evol. Microbiol.Home page
S. Yoshizawa, M. Wada, K. Kita-Tsukamoto, E. Ikemoto, A. Yokota, and K. Kogure
Vibrio azureus sp. nov., a luminous marine bacterium isolated from seawater
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1645 - 1649.
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Int. J. Syst. Evol. Microbiol.Home page
Y.-G. Chen, Y.-Q. Zhang, J.-X. Shi, H.-D. Xiao, S.-K. Tang, Z.-X. Liu, K. Huang, X.-L. Cui, and W.-J. Li
Jeotgalicoccus marinus sp. nov., a marine bacterium isolated from a sea urchin
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1625 - 1629.
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Int. J. Syst. Evol. Microbiol.Home page
P. H. Summanen, P. A. Lawson, and S. M. Finegold
Porphyromonas bennonis sp. nov., isolated from human clinical specimens
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1727 - 1732.
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Int. J. Syst. Evol. Microbiol.Home page
H.-Y. Weon, R. Anandham, B.-Y. Kim, S.-B. Hong, Y.-A. Jeon, and S.-W. Kwon
Dyella soli sp. nov. and Dyella terrae sp. nov., isolated from soil
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1685 - 1690.
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Int. J. Syst. Evol. Microbiol.Home page
L.-P. Zhang, L.-M. Zhang, and X.-M. Zhang
Streptosporangium canum sp. nov., isolated from soil
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1715 - 1719.
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Int. J. Syst. Evol. Microbiol.Home page
N. S. Gordon, A. Valenzuela, S. M. Adams, P. W. Ramsey, J. L. Pollock, W. E. Holben, and J. E. Gannon
Pedobacter nyackensis sp. nov., Pedobacter alluvionis sp. nov. and Pedobacter borealis sp. nov., isolated from Montana flood-plain sediment and forest soil
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1720 - 1726.
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Int. J. Syst. Evol. Microbiol.Home page
Y.-G. Chen, Y.-Q. Zhang, H.-D. Xiao, Z.-X. Liu, L.-B. Yi, J.-X. Shi, X.-Y. Zhi, X.-L. Cui, and W.-J. Li
Pontibacillus halophilus sp. nov., a moderately halophilic bacterium isolated from a sea urchin
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1635 - 1639.
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Int. J. Syst. Evol. Microbiol.Home page
T. Clavel, C. Charrier, A. Braune, M. Wenning, M. Blaut, and D. Haller
Isolation of bacteria from the ileal mucosa of TNFdeltaARE mice and description of Enterorhabdus mucosicola gen. nov., sp. nov.
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1805 - 1812.
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Int. J. Syst. Evol. Microbiol.Home page
L. Zhang, Y. Wang, J. Dai, Y. Tang, Q. Yang, X. Luo, and C. Fang
Bacillus korlensis sp. nov., a moderately halotolerant bacterium isolated from a sand soil sample in China
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1787 - 1792.
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Int. J. Syst. Evol. Microbiol.Home page
I. Cleenwerck, M. De Wachter, A. Gonzalez, L. De Vuyst, and P. De Vos
Differentiation of species of the family Acetobacteraceae by AFLP DNA fingerprinting: Gluconacetobacter kombuchae is a later heterotypic synonym of Gluconacetobacter hansenii
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1771 - 1786.
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Int. J. Syst. Evol. Microbiol.Home page
S.-K. Tang, Y. Wang, K. Lou, P.-H. Mao, X. Jin, C.-L. Jiang, L.-H. Xu, and W.-J. Li
Gracilibacillus saliphilus sp. nov., a moderately halophilic bacterium isolated from a salt lake
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1620 - 1624.
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Int. J. Syst. Evol. Microbiol.Home page
F. Peng, M. Liu, L. Zhang, J. Dai, X. Luo, H. An, and C. Fang
Planobacterium taklimakanense gen. nov., sp. nov., a member of the family Flavobacteriaceae that exhibits swimming motility, isolated from desert soil
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1672 - 1678.
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Int. J. Syst. Evol. Microbiol.Home page
H.-Y. Weon, B.-Y. Kim, S.-B. Hong, Y.-A. Jeon, B.-S. Koo, S.-W. Kwon, and E. Stackebrandt
Massilia niabensis sp. nov. and Massilia niastensis sp. nov., isolated from air samples
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1656 - 1660.
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Int. J. Syst. Evol. Microbiol.Home page
J.-H. Kim, K.-Y. Kim, and C.-J. Cha
Flavobacterium chungangense sp. nov., isolated from a freshwater lake
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1754 - 1758.
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Int. J. Syst. Evol. Microbiol.Home page
P. Schumann, P. Kampfer, H.-J. Busse, L. I. Evtushenko, and for the Subcommittee on the Taxonomy of the Subord
Proposed minimal standards for describing new genera and species of the suborder Micrococcineae
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1823 - 1849.
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Int. J. Syst. Evol. Microbiol.Home page
E. V. Pikuta, R. B. Hoover, A. K. Bej, D. Marsic, W. B. Whitman, and P. Krader
Spirochaeta dissipatitropha sp. nov., an alkaliphilic, obligately anaerobic bacterium, and emended description of the genus Spirochaeta Ehrenberg 1835
Int J Syst Evol Microbiol, July 1, 2009; 59(7): 1798 - 1804.
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Appl. Environ. Microbiol.Home page
Y.-B. Ahn, J.-C. Chae, G. J. Zylstra, and M. M. Haggblom
Degradation of Phenol via Phenylphosphate and Carboxylation to 4-Hydroxybenzoate by a Newly Isolated Strain of the Sulfate-Reducing Bacterium Desulfobacterium anilini
Appl. Envir. Microbiol., July 1, 2009; 75(13): 4248 - 4253.
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Int. J. Syst. Evol. Microbiol.Home page
H.-Y. Weon, S.-H. Yoo, Y.-J. Kim, J.-A Son, B.-Y. Kim, S.-W. Kwon, and B.-S. Koo
Chitinophaga niabensis sp. nov. and Chitinophaga niastensis sp. nov., isolated from soil
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1267 - 1271.
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Int. J. Syst. Evol. Microbiol.Home page
H.-Y. Weon, S.-H. Yoo, B.-Y. Kim, J.-A Son, Y.-J. Kim, and S.-W. Kwon
Niabella ginsengisoli sp. nov., isolated from soil cultivated with Korean ginseng
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1282 - 1285.
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Int. J. Syst. Evol. Microbiol.Home page
M. Liu, X. Luo, L. Zhang, J. Dai, Y. Wang, Y. Tang, J. Li, T. Sun, and C. Fang
Pseudomonas xinjiangensis sp. nov., a moderately thermotolerant bacterium isolated from desert sand
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1286 - 1289.
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Int. J. Syst. Evol. Microbiol.Home page
T.-W. Guan, S.-K. Tang, J.-Y. Wu, X.-Y. Zhi, L.-H. Xu, L.-L. Zhang, and W.-J. Li
Haloglycomyces albus gen. nov., sp. nov., a halophilic, filamentous actinomycete of the family Glycomycetaceae
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1297 - 1301.
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Int. J. Syst. Evol. Microbiol.Home page
K. Alain, N. Callac, M. Guegan, F. Lesongeur, P. Crassous, M.-A. Cambon-Bonavita, J. Querellou, and D. Prieur
Nautilia abyssi sp. nov., a thermophilic, chemolithoautotrophic, sulfur-reducing bacterium isolated from an East Pacific Rise hydrothermal vent
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1310 - 1315.
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Int. J. Syst. Evol. Microbiol.Home page
S.-K. Tang, Y. Wang, K. Lou, P.-H. Mao, L.-H. Xu, C.-L. Jiang, C.-J. Kim, and W.-J. Li
Kocuria halotolerans sp. nov., an actinobacterium isolated from a saline soil in China
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1316 - 1320.
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Int. J. Syst. Evol. Microbiol.Home page
Y.-G. Chen, X.-L. Cui, Q.-Y. Li, Y.-X. Wang, S.-K. Tang, Z.-X. Liu, M.-L. Wen, Q. Peng, and L.-H. Xu
Saccharospirillum salsuginis sp. nov., a gammaproteobacterium from a subterranean brine
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1382 - 1386.
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Int. J. Syst. Evol. Microbiol.Home page
L. Collado, I. Cleenwerck, S. Van Trappen, P. De Vos, and M. J. Figueras
Arcobacter mytili sp. nov., an indoxyl acetate-hydrolysis-negative bacterium isolated from mussels
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1391 - 1396.
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Int. J. Syst. Evol. Microbiol.Home page
D. W. Lee and S. D. Lee
Dyella marensis sp. nov., isolated from cliff soil
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1397 - 1400.
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Int. J. Syst. Evol. Microbiol.Home page
S. D. Lee
Amycolatopsis ultiminotia sp. nov., isolated from rhizosphere soil, and emended description of the genus Amycolatopsis
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1401 - 1404.
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Int. J. Syst. Evol. Microbiol.Home page
D. Wurdemann, B. J. Tindall, R. Pukall, H. Lunsdorf, C. Strompl, T. Namuth, H. Nahrstedt, M. Wos-Oxley, S. Ott, S. Schreiber, et al.
Gordonibacter pamelaeae gen. nov., sp. nov., a new member of the Coriobacteriaceae isolated from a patient with Crohn's disease, and reclassification of Eggerthella hongkongensis Lau et al. 2006 as Paraeggerthella hongkongensis gen. nov., comb. nov.
Int J Syst Evol Microbiol, June 1, 2009; 59(6): 1405 - 1415.
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