Selection of Nucleic Acid Aptamers Specific for Mycobacterium tuberculosis
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Tuberculosis (TB) remains to be a major global health problem, with about 9 million new cases and 1.4 million deaths in 2011. For the control of tuberculosis as well as other infectious diseases, WHO recommended “ASSURED” (Affordable, Sensitive, Specific, User-friendly, Rapid and robust, Equipment-free, and Deliverable to the end user) diagnostic tools that can easily be maintained and used in developing countries. Aptamers are promising tools for developing point-of-care diagnostic assays for TB. In this study, ssDNA aptamers that recognize Mycobacterium tuberculosis H37Ra were selected by systematic evolution of ligands by exponential enrichment (SELEX). For this purpose, two different selection protocols, ultrafiltration and centrifugation, were applied. A total of 21 TB specific aptamers were selected. These aptamers exhibited “G-rich” regions on the 3′ terminus of the aptamers, including a motif of “TGGGG,” “GTGG,” or “CTGG.” Binding capability of selected aptamers were investigated by quantitative PCR and Mtb36 DNA aptamer was found the most specific aptamer to M. tuberculosis H37Ra. The dissociation constant (K d) of Mtb36 aptamer was calculated as 5.09 ± 1.43 nM in 95 % confidence interval. Relative binding ratio of Mtb36 aptamer to M. tuberculosis H37Ra over Mycobacterium bovis and Escherichia coli was also determined about 4 times and 70 times more, respectively. Mtb36 aptamer is highly selective for M. tuberculosis, and it can be used in an aptamer-based biosensor for the detection of M. tuberculosis.
KeywordsTuberculosis Diagnosis Oligonucleotide Aptamer Selection SELEX Biosensor
This study was supported by The Scientific and Technological Research Council of Türkiye (TÜBİTAK) National Metrology Institute with project number 110S141. We would like to thank Prof. Dr. Asım Esen for his molecular biology expertise and assistance.
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