In Vitro Synthesis of Cas9 mRNA and gRNA

The in vitro synthesis of Cas9 mRNA and gRNA was performed as described previously^{8 (link)} using T3 RNA polymerase (New England Biolabs, MA, USA). The RNA transcripts were precipitated with absolute ethanol, washed, and resuspended in RNase-free water. The RNA solutions were stored at −80 °C until use.

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Fujii W., Ito H., Kanke T., Ikeda A., Sugiura K, & Naito K. (2019). Generation of genetically modified mice using SpCas9-NG engineered nuclease. Scientific Reports, 9, 12878.

Publication 2019

T3 RNA polymerase

Absolute ethanol Mrna Rnase Synthesis T3 rna polymerase

Corresponding Organization : The University of Tokyo

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Variable analysis

independent variables

In vitro synthesis of Cas9 mRNA and gRNA

dependent variables

Not explicitly mentioned

control variables

T3 RNA polymerase (New England Biolabs, MA, USA) used for in vitro synthesis of Cas9 mRNA and gRNA
Precipitation of RNA transcripts with absolute ethanol, washing, and resuspension in RNase-free water
Storage of RNA solutions at -80 °C until use

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

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