A modified T-vector for simplified assembly of hairpin RNAi constructs
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We describe a modified T-vector, pGFPm-T, for direct cloning of RT-PCR products to generate bidirectional restriction fragments for assembly of hairpin-containing RNAi vectors in the popular pFGC and pGSA binary vector backbone. Green fluorescence protein (GFP) is used as a visual reporter for direct selection of recombinants under UV illumination. The simplified cloning process enables a seamless workflow from candidate gene selection and RT-PCR verification to inverted repeat cloning, using a single pair of gene-specific primers.
KeywordsCloning vector Dihydroflavonol reductase genes Gene silencing GFP Populus Transgenic plant
We thank Dr. Soon-Yong Choi (HanNam University, South Korea) for providing the pHNT vector. This work was supported by the US Department of Energy, Office of Science, Biological and Environmental Research Program (DE-FG02-05ER64112), and the US National Science Foundation Plant Genome Program (DBI-0421756).
- Park KS, Park SW, Choi SY (2000) Improved T-vector for the cloning of PCR DNA using green fluorescent protein. J Microbiol Biotechnol 10:264–266Google Scholar