Changlin Fu

Publications (2)12.12 Total impact

• Source

  Available from: PubMed Central

  Article: Single tube, high throughput cloning of inverted repeat constructs for double-stranded RNA expression.

  Brian Hauge, Christopher Oggero, Nicole Nguyen, Changlin Fu, Fenggao Dong
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  ABSTRACT: RNA interference (RNAi) has emerged as a powerful tool for the targeted knockout of genes for functional genomics, system biology studies and drug discovery applications. To meet the requirements for high throughput screening in plants we have developed a new method for the rapid assembly of inverted repeat-containing constructs for the in vivo production of dsRNAs. The procedure that we describe is based on tagging the sense and antisense fragments with unique single-stranded (ss) tails which are then assembled in a single tube Ligase Independent Cloning (LIC) reaction. Since the assembly reaction is based on the annealing of unique complementary single stranded tails which can only assemble in one orientation, greater than ninety percent of the resultant clones contain the desired insert. Our single-tube reaction provides a highly efficient method for the assembly of inverted repeat constructs for gene suppression applications. The single tube assembly is directional, highly efficient and readily adapted for high throughput applications.
  PLoS ONE 01/2009; 4(9):e7205. · 4.09 Impact Factor
• Source

  Available from: PubMed Central

  Article: Rapid one-step recombinational cloning.

  Changlin Fu, Daniel R Wehr, Janice Edwards, Brian Hauge
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  ABSTRACT: As an increasing number of genes and open reading frames of unknown function are discovered, expression of the encoded proteins is critical toward establishing function. Accordingly, there is an increased need for highly efficient, high-fidelity methods for directional cloning. Among the available methods, site-specific recombination-based cloning techniques, which eliminate the use of restriction endonucleases and ligase, have been widely used for high-throughput (HTP) procedures. We have developed a recombination cloning method, which uses truncated recombination sites to clone PCR products directly into destination/expression vectors, thereby bypassing the requirement for first producing an entry clone. Cloning efficiencies in excess of 80% are obtained providing a highly efficient method for directional HTP cloning.
  Nucleic Acids Research 06/2008; 36(9):e54. · 8.03 Impact Factor