Journal
METHODS
Volume 30, Issue 4, Pages 289-295Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/S1046-2023(03)00036-7
Keywords
gene silencing; recombination cloning; plant functional genomics; pHELLSGATE
Ask authors/readers for more resources
Gene silencing can be achieved by transformation of plants with constructs that express self-complementary (termed hairpin) RNA containing sequences homologous to the target genes. The DNA sequences encoding the self-complementary regions of hairpin (hp) RNA constructs form an inverted repeat. The inverted repeat can be stabilized in bacteria through separation of the self-complementary regions by a spacer region. When the spacer sequence encodes an intron, the efficiency of gene silencing is very high. There are at least three ways in which hpRNA constructs can be made. The construct may be generated from standard binary plant transformation vectors in which the hairpin-encoding region is generated de novo for each gene. Alternatively, generic gene-silencing vectors such as the pHANNIBAL and the pHELLSGATE series can be used. They simply require the insertion of PCR products, derived from the target gene, into the vectors by conventional cloning or by using the Gateway directed recombination system. In this article, we describe and evaluate the advantages of these vectors and then provide the protocols for their efficient use. (C) 2003 Published by Elsevier Science (USA).
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available