geoscience.net logo
+ Resolve Article
+ Follow Us
Follow on FacebookFollow on Facebook
Follow on TwitterFollow on Twitter

+ Translate
+ Subscribe to Site Feed
GeoScience Most Shared ContentMost Shared Content

Label-free selection of RNA aptamers for metabolic engineering


, : Label-free selection of RNA aptamers for metabolic engineering. Methods 106: 37-41

RNA aptamers can be assembled into genetic regulatory devices that sense and respond to levels of specific cellular metabolites and thus serve an integral part of designing dynamic control into engineered metabolic pathways. Here, we describe a practical method for generating specific and high affinity aptamers to enable the wider use of in vitro selection and a broader application of aptamers for metabolic engineering. Conventional selection methods involving either radioactive labeling of RNA or the use of label-free methods such as SPR to track aptamer enrichment require resources that are not widely accessible to research groups. We present a label-free selection method that uses small volume spectrophotometers to track RNA enrichment paired with previously characterized affinity chromatography methods. Borrowing techniques used in solid phase peptide synthesis, we present an approach for immobilizing a wide range of metabolites to an amino PEGA matrix. As an illustration, we detail laboratory techniques employed to generate aptamers that bind p-aminophenylalanine, a metabolic precursor for bio-based production of plastics and the pristinamycin family of antibiotics. We focused on the development of methods for ligand immobilization, selection via affinity chromatography, and nucleic acid quantification that can be performed with common laboratory equipment.

US$19.90

PMID: 27339940

DOI: 10.1016/j.ymeth.2016.06.016


Other references

Capellini, G., 1893: Nuovi Resti di Zifioidi in Calabria e in Toscana. RendLincei: 283-288

Sharma, G.V.M.; Chandramouli, N.; Basha, S.Jeelani.; Nagendar, P.; Ramakrishna, K.V.S.; Sarma, A.V.S., 2011: The design of α/β-peptides: study on three-residue turn motifs and the influence of achiral glycine on helix and turn. Novel three-residue helix-turn secondary structures, nucleated by a helix at the N terminus, were generated in peptides that have 'β-Caa-L-Ala-L-Ala,' 'β-Caa-L-Ala-γ-Caa,' and 'β-Caa-L-Ala-δ-Caa' (in which β-Caa...

Lewandowski Ann M., 2008: Costs of groundwater nitrate contamination. Journal of Soil and Water Conservation 63(3): 92A

D'haens, G.; Bourgeois, S.; Hiele, M.; Van Assche, G.; Lemmens, L.; Rutgeerts, P., 2000: Two and four mg/kg/day of IV cyclosporine are equally effective in severe attacks of ulcerative colitis. Digestive & Liver Disease 32(Supplement 1): A18, May

Re, P., 1979: The eggs and planktonic stages of Portuguese marine fishes. 2. - Ichthyoplankton of Tejo Estuary (1978). A preliminary survey of fish eggs and larvae occurring in Tejo estuary was made during 1978. Sampling was carried at 7 stations with a 2 m ring trawl of 0.300 mm mesh aperture. Successive oblique trawls (3) were made at each station for 10 min per...

Evrard, E.; Molders, V.; D.B.Uyne, M.Tta, D., 1960: Contribution to the study of R-1625 or haloperidol in chronic psychoses or behavior disorders of certain mentally degenerated patients. Acta Neurologica et Psychiatrica Belgica 60: 811-823

Mahnken, A.H.; Brandenburg, V.M.; Frank, R.D.; Haage, P.; Günther, R.W., 2004: Hemolysis, hemorrhage, headache, and hidden abortion: imaging findings in antiphospholipid syndrome. Antiphospholipid antibodies are associated with arterial and venous thromboses, recurrent pregnancy loss, and organ infarction. Any vascular region can be affected. We present a 20-year-old woman suffering from secondary antiphospholipid syndrome...

Mtwisha, L.B.andt, W.M.cready, S.L.ndsey, G., 1998: HSP 12 is a LEA-like protein in Saccharomyces cerevisiae. LEA group I, II and III antibodies all recognised soluble proteins present in an extract of yeast (Saccharomyces cerevisiae). The smaller protein of the two recognised by the group I antibody displayed identical migration on SDS-PAGE to the pea se...

Kusumi, A.; Subczynski, W.K.; Hyde, J.S., 1982: Oxygen transport parameter in membranes as deduced by saturation recovery measurements of spin-lattice relaxation times of spin labels. Spin-lattice relaxation time (T1) measurements of nitroxide radical spin labels in membranes have been made by using the saturation-recovery technique. Stearic acid and sterol-type labels were used as probes of dimyristoylphosphatidylcholine lipos...

Ham, P.Benson.; Hwang, B.; Wise, L.J.; Walters, K.Christian.; Pipkin, W.L.; Howell, C.G.; Bhatia, J.; Hatley, R., 2016: Venovenous Extracorporeal Membrane Oxygenation in Pediatric Respiratory Failure. Conventional treatment of respiratory failure involves positive pressure ventilation that can worsen lung damage. Extracorporeal membrane oxygenation (ECMO) is typically used when conventional therapy fails. In this study, we evaluated the use of...