title: [Screening and structure analysis of the aptamer target to Escherichia coli tolC protein].
abstract: OBJECTIVE
  To screen and characterize the aptamer of Escherichia coli outer member protein tolC.
  
  
  METHODS
  By using the recombinant E.coli outer member protein tolC for the screening target, oligonucleotides which were capable of specifically binding to the protein were screened from a random oligonucleotide library through the stematic evolution of ligand by exponential enrichment (SELEX) technique. The binding capacity of ssDNA to the targeted protein from each round was detected by the FITC fluorescence labeling technique.The ssDNA from the last cycle was cloned and sequenced,and the second structure was further analyzed by the DNAMan program.
  
  
  RESULTS
  After 12 cycles of selection, 40 clones were selected randomly and sequenced. Although a unique conserved sequence was not obtained among the 23 obtained aptamers by the primary structure analysis,three pairs of aptamers and two pairs of aptamers were found to be identical.Analysis of the secondary structure revealed that the stem-loop and bulge loop were the main motifs,indicating that they might play a key role in the binding of aptamers to the target protein. According to the characteristic of the second structure,23 aptamers were divided into four families,and aptamer 20 bore the greatest affinity.
  
  
  CONCLUSIONS
  Aptamers against E.coli outer member protein tolC were successfully identified by the SELEX method. The results laid a foundation for the investigation of the interference to the drug resistance of E. coli and the underlying mechanisms.
authors.affiliation: Medical school of Hunan University of Chinese Medicine, Changsha 410208, China.
authors.forename: Ling-li
authors.initials: LL
authors.lastname: Chen
authors.affiliation: Medical school of Hunan University of Chinese Medicine, Changsha 410208, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Li
authors.affiliation: Medical school of Hunan University of Chinese Medicine, Changsha 410208, China.
authors.forename: Xiao-qing
authors.initials: XQ
authors.lastname: Zhang
authors.affiliation: Medical school of Hunan University of Chinese Medicine, Changsha 410208, China.
authors.forename: Lan
authors.initials: L
authors.lastname: Song
authors.affiliation: Medical school of Hunan University of Chinese Medicine, Changsha 410208, China.
authors.forename: Chun
authors.initials: C
authors.lastname: Qian
authors.affiliation: Medical school of Hunan University of Chinese Medicine, Changsha 410208, China.
authors.forename: Ji-wen
authors.initials: JW
authors.lastname: Ge
mesh_terms: D052157:Aptamers, Nucleotide / Q000235:genetics*; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry*; D004277:DNA, Single-Stranded; D004926:Escherichia coli; D029968:Escherichia coli Proteins / Q000737:chemistry*; D015723:Gene Library; D026901:Membrane Transport Proteins / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D052156:SELEX Aptamer Technique
chemical_list: D052157:Aptamers, Nucleotide; D001425:Bacterial Outer Membrane Proteins; D004277:DNA, Single-Stranded; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D011994:Recombinant Proteins; C073126:tolC protein, E coli