Experiment Validating Gene Editing Reporter 14 (GER14) Mouse Model in Heterozygous Blastocysts Experiment Condition GER14-g1-SpC9-FWD Assay Description: Reporter activation by NHEJ is calculated as the percent of blasts that express Katushka2S (RFP) out of the total blasts imaged. Tissue Measured: Measurement Type: Editing Efficiency Record ID 15000003591 Editor Alt-R™ S.p. Cas9 Nuclease V3 Delivery System Delivery System Subtype Guide GER14-g1-SpC9-FWD Vector Model Species Mouse Model Name GER14 Heterozygous Blastocyst Application Method Electroporation Application Site Dosage Injection Frequency Injection Rate Injection Volume Days post injection 96 hours Editor Format ribonucleoprotein (RNP) Antidote Id Antidote Description Measured Values Samples Size: 0   Replicate Result (Editing Efficiency) Measurement Units: % of blasts with RFP activation Mean 59.48  1 60.42  2 46.15  3 71.88  Measurement Units: % of blasts with edits Mean 98.85  1 100  2 100  3 96.55  Measurement Units: Signal Mean # blasts analyzed  1 93 blasts for RFP  Mean # blasts analyzed  1 90 blasts for edits

SpCas9 Validation of Gene Editing Reporter 14 (GER14) Mouse Model in Heterozygous Blastocysts Zygotes were electroporated with SpCas9 RNP along with guides and cultured 96 hours to blastocyst stage. Blastocysts were imaged for Katushka2S (RFP) fluorescence and scored for fluorescence signal.