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. 2015 Sep;21(9):1683–1689. doi: 10.1261/rna.051631.115

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© 2015 Mefferd et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society

This article is distributed exclusively by the RNA Society for the first 12 months after the full-issue publication date (see http://rnajournal.cshlp.org/site/misc/terms.xhtml). After 12 months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.

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FIGURE 2. — Neisseria meningitidis sgRNA expression using tRNA-derived promoters. (A) Relative luciferase activity detected in 293T cells cotransfected with plasmids expressing Nme Cas9 and sgRNAs specific for protospacer 9 (P9) or 25 (P25) (Zhang et al. 2013) or a negative control sgRNA (Neg), and their cognate indicator plasmids. sgRNAs were expressed from either the U6 promoter or the indicated tRNA. Average of three experiments with SD indicated. (B) Northern blot of Nme sgRNAs recovered from 293T cells transfected with a plasmid expressing Nme Cas9 and sgRNAs specific for the P25 sequence expressed using U6 or the indicated tRNA. Endogenous cellular U6 RNA served as a loading control. (C) Same as B except that expression of a variety of distinct sgRNA guide sequences, specific for the indicated gene sequence, was analyzed. (P25) protospacer 25; (P9) protospacer 9; (ICP4) HSV-1 infected cell protein 4; (GFP) green fluorescent protein; (G6P) glucose-6-phosphatase.