miRZips anti-microRNAs
Permanent microRNA suppression
Permanent microRNA interference
- Lentiviral-based anti-sense microRNAs
- Use both GFP and Puro selection
- Dissect microRNA regulated pathways
miRZip constructs express high levels of anti-microRNAs
MiRZip constructs were tested for anti-microRNA expression individually by transfecting HEK293 cells with 3ug miRZip plasmid DNA per well in 6 well tissue culture dishes using Lipofectamine 2000. The cells were lysed and total RNA extracted after 48 hours. The RNA samples were converted to cDNA using SBI's QuantiMir RT system for qPCR analysis. The levels of anti-microRNA expression were measured using QuantiMir primer assays designed for the specific miRZip anti-microRNA tested and compared to untransfected controls.
Expression validation of miRZip anti-microRNAs by qPCR
The QuantiMir cDNA from the transfections was diluted 1:50 and one microliter was used as template in 6ul qPCR reactions along with the primer assay for the specific anti-microRNA sequence labeled below. Standard thermal cycling conditions were used and data graphed as miRZip expression level compared to untransfected controls.
