Silenciamiento de la fosfoglucosa isomerasa (PGI) en fibroblastos inmortalizados de ratón. Mediante la técnica del RNA de interferencia

Abstract

A target-specific small hairpin RNA (shRNA) was added into a plasmid delivery system (pSUPER-NeoGFP) and introduced into a cell culture of mouse embryonic fibroblasts (MEFs). The shRNA was constructed for suppressing   the expression in mice cells glucose-6-phosphate isomerase PGIm. Glucose-6-phosphate isomerase is an enzyme that catalyses the conversion of glucose-6-phosphate into fructose 6-phosphate in glycolysis’ second step. A PEI–polyethylene glycol conjugate was used as a pSUPER-NeoGFP-shPGI plasmid carrier. Two cultures of cells were transformed one used as control with the plasmid delivery system (pSUPER-NeoGFP) and the other with the plasmid with the sh-PGI. The gene inhibition and cellular uptake behaviours were explored by the measurement of lactic acid as a marker of the efficiency of glycolysis and with a reverse transcription polymerase chain reaction (RT-PCR) to quantify PGIm expresion. No significatively differences were found in the lactic acid production while a decrease in the PGIm expresion was observed.