Knockout of Micro-RNA-21 Gene in DLBCL OCI-Ly3 cells by TALEN Technique / 中国实验血液学杂志

ABSTRACT

<p><b>OBJECTIVE</b>To explore an efficient way to knockout microRNA genes in hemapoietic cell lines with a very low transfection efficiency, so as to facilitate the study of microRNA function in hematopoietic malignancies.</p><p><b>METHODS</b>TALE-nucleases was utilized to knockout the microRNA-21 gene in human diffuse large B-cell lymphoma cells (OCI-Ly3). The OCI-Ly3 single cell clones without expression of miR-21 were established through eGFP(+) enrichment, PCR screening, and microRNA quantification. Finally, the miR-21 changes of mutant clones were identified by sequencing.</p><p><b>RESULTS</b>Four miR-21-knockouted OCI-Ly3 single-cell-derived clones were established after 2 round transfection and screening. The miR-21 knockout efficiency was around 10/10(6) original cells. Sequencing the mutant clones indicated that miR-21 expression could be drastically reduced by simply altering sequences immediately adjacent to the microRNA duplex.</p><p><b>CONCLUSION</b>This strategy may be applied to knockout any microRNA of interest even in hemapoietic cell lines with very low transfection efficiency.</p>