A strategy to reduce miRNAs activity in plants based on artificial miRNAs

MANACORDA, CA; BAZZINI, AA; CONTI, G; ASURMENDI, S

San Miguel de Tucumán - Prov. Tucumán - Argentina

Congreso; XLV Reunión Anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2009

Sociedad Argentina de Investigación Bioquímica

MicroRNAs (miRNAs) are 19-24-nucleotide ssRNA molecules that regulate gene expression and play a key role in plant development and morphogenesis. They are usually multigenic families whose members are expressed in a temporally and spatially regulated pattern with partially overlapping functions. miR164 family (a, b and c) regulate a group of NAC-domain containing transcription factors required for plant development. MiRNAs genetics knock out are useful but laborious and time-consuming gen-by-gen approaches. In this work, we present a novel tool to reduce the activity of the entire miR164 family. An artificial miRNA designed to recognize mature miR164 was cloned under the 35S promoter and transgenic Arabidopsis thaliana plants were generated. Transgenic expression of the artificial miRNA was confirmed by qRT-PCR in several lines.Phenotypic analysis of T1 lines showed morphological alterations that resemble those observed in miR164abc loss-of-function triple mutants. However, the severity of the phenotypes was attenuated gradually in further self-crossed generations. Northern blot analysis revealed changes in the expression level of endogenous miR164 in some transgenic lines. This approach could be a valuable tool, to study miRNAs functions as well as a to use it biotechnologically. Although epigenetic regulation of the artificial miRNA might be playing a role limiting its usefulness.