title: First step in pre-miRNAs processing by human Dicer.
abstract: OBJECTIVE
  To investigate the strand preference of the initial cleavage of human pre-miRNAs by human Dicer in vitro.
  
  
  METHODS
  We used a series of in vitro transcribed pre-miRNAs that were radioactively labeled at their 5' or 3' ends in cleavage reactions with recombinant human Dicer or HeLa cytoplasmic S100 extracts. Pre-miRNAs samples were purified by denaturing and native PAGE and only the stem-loop structures were used in the experiments. Products of cleavage reactions were resolved by denaturing PAGE, and scanned by phosphor-imaging.
  
  
  RESULTS
  Recombinant hDicer performs a biased first-cleavage in the pre-let-7b and hsa-pre-miR-17 3' strand. This result is recapitulated in HeLa S100 cytoplasmic extracts.
  
  
  CONCLUSIONS
  The differential first-nick is observed in cleavage reactions only when stem-loops are substrates for hDicer.
authors: Departamento de Biología Celular, Instituto de Fisiología Celular, UNAM Apartado Postal 70-600, Circuito exterior s/n Ciudad Universitaria, Delegación Coyoacan, CP 04510, México DF.
authors: Carlos Fabián
authors: CF
authors: Flores-Jasso
authors: Catalina
authors: C
authors: Arenas-Huertero
authors: Jose Luis
authors: JL
authors: Reyes
authors: Cecilia
authors: C
authors: Contreras-Cubas
authors: Alejandra
authors: A
authors: Covarrubias
authors: Luis
authors: L
authors: Vaca
mesh_terms: D001483:Base Sequence; D006367:HeLa Cells; D006801:Humans; D035683:MicroRNAs / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D043244:Ribonuclease III / Q000235:genetics / Q000378:metabolism*
chemical_list: D035683:MicroRNAs; D011994:Recombinant Proteins; D043244:Ribonuclease III