Figure 5.

Evaluation of siMVP cytotoxicity. (A) siMVP-containing plasmids were isolated either directly from the library (selection cycle 0), or from cells that survived in each selection cycle with 150 μM 5FC, or from dead cells resulted in the first cycle of bacterial invasion and drug selection and transformed into E. coli. For toxicity evaluation, 30 colonies were randomly selected from each transformation for reisolation of siMVP-containing plasmids. HeLa cells in 96-well plate transfected in triplicate with siMVP-containing plasmids (0.2 μg) prepared from individual colonies. (B) HeLa cells transfected with 10 nM of siRNAs synthesized in vitro. Cell survival values represent the mean of three independent transfections each performed in triplicate. Five highly toxic siMVPs, siMVP-28, siMVP-34, siMVP-52, siMVP-59 and siMVP-61, were identified from dead cells (A) after screening the transformed100 colonies. Their toxicity was compared with most toxic SRD5a2-3 reported. Cellular viability assays in both (A) and (B) were performed at 72-h after transfection. The dotted line represents the 75% cell viability threshold. Data in (A) and (B) are mean ± SD.