Abstract
Streptomycin-resistantMycobacterium tuberculosis has been attributed to two distinct classes of mutations, including point mutations within therpsL gene (three mutation sites) and therrs gene (seven mutation sites). We have developed an automated simultaneous detection system of multiple mutations based on thermal dissociation curve analysis for streptomycin resistance inM. tuberculosis using streptavidin-labeled bacterial magnetic particles (SA-BacMPs). With consideration for time and cost effectiveness, we used fewer PCR reactions, with a long PCR target (rpsL, 182 bp;rrs, 467 bp) including multiple mutation sites. In order to improve the amount of target DNA captured on BacMPs through streptavidin-biotin binding, several reaction conditions, such as salt species and concentration in the buffer, and reaction temperature were examined. Compared to the commonly used 1M NaCl solution, the amount of DNA captured on SA-BacMPs was about six times greater (approx 5 pmoles/50 μg BacMPs) in the 2M LiCl solution. Under these conditions, automated nucleotide discriminations of 10 targets inrpsL andrrs genes of streptomycin-resistant and wild-type strains were successfully performed at the same time.
Similar content being viewed by others
References
Phadtare, S., Vinod, V. P., Wadgaonkar, P. P., Rao, M., and Sastry, M. (2004),Langmuir 20, 3717–3723.
Yang, H. H., Zhang, S. Q., Chen, X. L., Zhuang, Z. X., Xu, J. G., and Wang, X. R. (2004),Anal. Chem. 76, 1316–1321.
Akerman, M. E., Chan, W. C., Laakkonen, P., Bhatia, S. N., and Ruoslahti, E. (2002),Proc. Natl. Acad. Sci. USA 99, 12,617–12,621.
Gao, X., Cui, Y., Levenson, R. M., Chung, L. W., and Nie, S. (2004),Nat. Biotechnol. 22, 969–976.
Prasad, B. L., Stoeva, S. I., Sorensen, C. M., Zaikovski, V., and Klabunde, K. J. (2003),J. Am. Chem. Soc. 125, 10,488–10,489.
Bruckl, H., Panhorst, M., Schotter, J., Kamp, P. B., and Becker, A. (2005),IEE Proc. Nanobiotechnol. 152, 41–46.
Matsunaga, T., Tadokoro, F., and Nakamura, N. (1990).IEEE Trans. Magn. 26, 1557–1559.
Nakamura, N., Burgess, J. G., Yagiuda, K., Kudo, S., Sakaguchi, T., and Matsunaga, T. (1993).Anal. Chem. 65, 2036–2039.
Maruyama, K., Takeyama, H., Nemoto, E., Tanaka, T., Yoda, K., and Matsunaga, T. (2004),Biotechnol. Bioeng. 87, 687–694.
Tsuchiya, M., Nakao, H., Katoh, T., et al. (2005).Hum. Reprod. 20, 974–978.
Nakagawa, T., Hashimoto, R., Maruyama, K., Tanaka, T., Takeyama, H., and Matsunaga, T. (2006).Biotechnol. Bioeng. 94, 862–868.
Tong, X. and Smith, L. M. (1992),Anal. Chem. 64, 2672–2677.
Diaz-Gonzales, M., Gonzales-Garcia, M. B., and Costa-Garcia, A. (2005).Biosens. Bioelectron. 20, 2035–2043.
Pavlou, A. K., Magan, N., Jones, J. M., Brown, J., Klatser, P., and Turner, A. P. (2004).Biosens. Bioelectron. 20, 538–544.
Fend, R., Kolk, A. H., Bessant, C., Bujitels, P., Klatser, P. R., and Woodman, A. C. (2006).J. Clin. Microbiol. 44, 2039–2045.
Telenti, A. (1997),Clin. Chest Med. 18, 55–64.
Ramaswamy, S. and Musser, J. M. (1998),Tuber. Lung Dis. 79, 3–29.
Matsunaga, T., Kawasaki, M., Yu, X., Tsujimura, N., and Nakamura, N. (1996).Anal. Chem. 68, 3551–3554.
Tanaka, T., Maruyama, K., Yoda, K., et al. (2003),Biosens. Bioelectron. 19, 325–330.
Holmberg, A., Blomstergren, A., Nord, O., Lukacs, M., Lundeberg, J., and Uhlen, M. (2005),Electrophoresis 26, 501–510.
Brown, T. A. (1999),Genomes. Wiley-Liss, New York.
Kinoshita, M. and Harano, Y. Bull. (2005),Chem. Soc. Japan 78, 1431–1441.
Hyre, D. E., Amon, L. M., Penzotti, J. E., et al. (2002).Nat. Struct. Biol. 9, 582–585.
Collins, K. D. (1995),Proc. Natl. Acad. Sci. USA 92, 5553–5557.
Hribar, B., Southall, N. T., Vlachy, V., and Dill, K. A. (2002),J. Am. Chem. Soc. 124, 12,302–12,311.
Gorby, Y. A., Beveridge, T. J., and Blakemore, R. P. (1987).J. Bacteriol. 170, 834–841.
Tanaka, T. and Matsunaga, T. (2000),Anal. Chem. 72, 3518–3522.
Arakaki, A., Webb, J., and Matsunaga, T. (2003),J. Biol. Chem. 278, 8745–8750.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Maruyama, K., Uchida, N., Takeyama, H. et al. Simultaneous detection of multiple mutations conferring streptomycin resistance inMycobacterium tuberculosis using nanoscale engineered biomagnetites. Nanobiotechnol 2, 71–78 (2006). https://doi.org/10.1007/BF02697261
Issue Date:
DOI: https://doi.org/10.1007/BF02697261