Highly efficient gene silencing using perfect complementary artificial miRNA targeting AP1 or heteromeric artificial miRNA targeting AP1 and CAL genes.
Gene silencing is a useful technique for elucidating biological function of genes by knocking down their expression. Recently developed artificial microRNAs (amiRNAs) exploit an endogenous gene silencing mechanism that processes natural miRNA precursors to small silencing RNAs that target transcripts for degradation. Based on natural miRNA structures, amiRNAs are commonly designed such that they have a few mismatching nucleotides with respect to their target sites as well as within mature amiRNA duplexes. In this study, we performed an analysis in which the conventional and modified form of an amiRNA was compared side by side. We showed that the amiRNA containing 5' mismatch with its amiRNA* and perfect complementarity to its target gene acted as a highly potent gene silencing agent against AP1, achieving a desired null mutation effect. In addition, a simultaneous silencing of two independent genes, AP1 and CAL1 was tested by employing a multimeric form of amiRNAs. Advantages and potential disadvantages of using amiRNAs with perfect complementarity to the target gene are discussed. The results presented here should be helpful in designing more specific and effective gene silencing agents.
Park
Wonkeun
W
Department of Plant and Soil Sciences, Delaware Biotechnology Institute, University of Delaware, Newark, DE 19711, USA.
Zhai
Jixian
J
Lee
Jung-Youn
JY
D017360:Arabidopsis / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics*; D018744:DNA, Plant / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D017343:Genes, Plant; D026161:MADS Domain Proteins / Q000235:genetics*; D035683:MicroRNAs / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000235:genetics
C091535:AP1 protein, Arabidopsis; D029681:Arabidopsis Proteins; C091536:CAL protein, Arabidopsis; D018744:DNA, Plant; D026161:MADS Domain Proteins; D035683:MicroRNAs