caa a22 4500 60550105X CHVBK 20210128100559.0 cr unu---uuuuu 210128e20151101xx s 000 0 eng 10.1007/s00253-015-6858-9 doi (NATIONALLICENCE)springer-10.1007/s00253-015-6858-9 A combinatorial systematic evolution of ligands by exponential enrichment method for selection of aptamer against protein targets [Elektronische Daten] [Bhairab Mondal, Shylaja Ramlal, Padma Lavu, Harishchandra Murali, Harsh Batra] Aptamers are synthetic DNA recognition elements which form unique conformations that enable them to bind specifically to their targets. In the present study, an attempt was made to standardize a new modified combinatorial method comprising of Ni-NTA affinity Systematic Evolution of Ligands by Exponential Enrichment (SELEX; based on affinity between His tag protein and Ni-NTA), membrane SELEX (based on immobilization of protein on nitrocellulose membrane), and microtiter plate based SELEX (to monitor affinity and to enrich the selected aptamers) for protein targets. For experimental evaluation, staphylococcal interotoxin B was the molecule chosen. The new combinatorial method enhanced selection ability up to 51.20% in comparison with individual conventional procedures. Employing this method following six rounds of selection, high-affinity aptamers with very different properties could be obtained with a dissociation constant (K d) value as low as 34.72 ± 25.09nM. The optimal aptamers could be employed in fluorescence binding assay, enzyme-linked oligonucleotide assays, and aptamer-based Western blot assay for characterization and detection. These results pave a potential path without using of any robotics for high-throughput generation of aptamers with advantages in terms of rapidity, simplicity, and ease in handling. Springer-Verlag Berlin Heidelberg, 2015 DNA aptamer nationallicence Combinatorial systematic evolution of ligands by exponential enrichment nationallicence Asymmetric polymerase chain reaction nationallicence Enterotoxin B nationallicence Enzyme-linked oligonucleotide assay nationallicence Mondal Bhairab Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut Ramlal Shylaja Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut Lavu Padma Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut Murali Harishchandra Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut Batra Harsh Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/22(2015-11-01), 9791-9803 0175-7598 99:22<9791 2015 99 253 https://doi.org/10.1007/s00253-015-6858-9 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6858-9 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Mondal Bhairab Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut NATIONALLICENCE 700 1- Ramlal Shylaja Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut NATIONALLICENCE 700 1- Lavu Padma Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut NATIONALLICENCE 700 1- Murali Harishchandra Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut NATIONALLICENCE 700 1- Batra Harsh Defence Food Research Laboratory (DFRL), 570011, Mysore, India aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/22(2015-11-01), 9791-9803 0175-7598 99:22<9791 2015 99 253