Correction: Gaussian decomposition of high-resolution melt curve derivatives for measuring genome-editing efficiency.

[This corrects the article DOI: 10.1371/journal.pone.

Gaussian decomposition of high-resolution melt curve derivatives for measuring genome-editing efficiency.

We describe a method for measuring genome editing efficiency from in silico analysis of high-resolution melt curve data. The melt curve data derived from amplicons of genome-edited or unmodified target sites were processed to remove the background fluorescent signal emanating from free fluorophore and then corrected for temperature-dependent quenching of fluorescence of double-stranded DNA-bound fluorophore. Corrected data were normalized and numerically differentiated to obtain the first derivatives of the melt curves.

Non-Homologous End Joining and Homology Directed DNA Repair Frequency of Double-Stranded Breaks Introduced by Genome Editing Reagents.

Genome editing using transcription-activator like effector nucleases or RNA guided nucleases allows one to precisely engineer desired changes within a given target sequence. The genome editing reagents introduce double stranded breaks (DSBs) at the target site which can then undergo DNA repair by non-homologous end joining (NHEJ) or homology directed recombination (HDR) when a template DNA molecule is available. NHEJ repair results in indel mutations at the target site.

Gammaretroviral vector encoding a fluorescent marker to facilitate detection of reprogrammed human fibroblasts during iPSC generation.

Induced pluripotent stem cells (iPSCs) are becoming mainstream tools to study mechanisms of development and disease. They have a broad range of applications in understanding disease processes, in vitro testing of novel therapies, and potential utility in regenerative medicine. Although the techniques for generating iPSCs are becoming more straightforward, scientists can expend considerable resources and time to establish this technology.

RRE-deleting self-inactivating and self-activating HIV-1 vectors for improved safety.

Retroviruses have been shown to efficiently delete sequences between repeats as a consequence of the template switching ability of the viral reverse transcriptase. To evaluate this approach for deriving safety-modified lentiviral vectors, we created HIV-1 vectors engineered to delete the Rev-response element (RRE) during reverse-transcription by sandwiching the RRE between two non-functional hygromycin phosphotransferase sequences. Deletion of the RRE during reverse-transcription lead to the reconstitution of a functional hygromycin phosphotransferase gene in the target cell.