A Novel ssDNA Aptamer Targeting Carcinoembryonic Antigen: Selection and Characterization
Yunussova N. Sypabekova M. Zhumabekova Z. Matkarimov B. Kanayeva D.
October 2022MDPI
Biology
2022#11Issue 10
One of the major causes of a drastically shorter life expectancy and one of the most prevalent diseases in the world today is cancer. Given the data on the rise in cancer cases throughout the world, it is obvious that, despite the diagnostic techniques currently being used, there is a pressing need to develop precise and sensitive techniques for early diagnosis of the disease. A high degree of affinity and specificity towards particular targets is maintained by the short nucleic acid molecules known as aptamers. Aptamers outperform antibodies due to their unique benefits, such as their simplicity in synthesis and modification, lack of toxicity, and long-term stability. Utilizing an accurate recognition element and a robust signal transduction mechanism, molecular diagnostics can be extremely sensitive and specific. In this study, development of new single-stranded DNA aptamers against CEA for use in cancer diagnostics was accomplished using SELEX and NGS methods. As a result of 12 iterative SELEX rounds, nine aptamer candidates against CEA were developed. NGS comparative analysis revealed that round twelve had an enriched number of aptamers that were specifically bound, as opposed to round eight. Among the selected nine sequences characterized by bioinformatics analysis and ELONA, an aptamer sequence with the highest specificity and affinity for the target protein was identified and further examined. Aptamer sequence (6) was screened in a concentration-dependent assay, specificity analysis was performed, and its potential secondary and tertiary structures were predicted, which enabled us to test one of the possible putative interactions with CEA. Finally, aptamer sequence (6) labelled with a Cy5 fluorescent tag was used in confocal microscopy to observe its binding towards the CEA expressed in HT-29 human colon adenocarcinoma cell line.
aptamer , bioinformatics analysis , carcinoembryonic antigen (CEA) , ELONA , HT-29 human colon adenocarcinoma , selection , SELEX
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Life Sciences, Nazarbayev University, 53 Kabanbay Batyr Ave, Astana, 010000, Kazakhstan
National Laboratory Astana, Nazarbayev University, 53 Kabanbay Batyr Ave, Astana, 010000, Kazakhstan
M.Sc. Program in Biological Sciences, Nazarbayev University, 53 Kabanbay Batyr Ave., Astana, 010000, Kazakhstan
Department of Biology, School of Sciences and Humanities, Nazarbayev University, 53 Kabanbay Batyr Ave., Astana, 010000, Kazakhstan
Life Sciences
National Laboratory Astana
M.Sc. Program in Biological Sciences
Department of Biology
10 лет помогаем публиковать статьи Международный издатель
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