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Project: The Jackson Laboratory Gene Editing Testing Center (JAX-GETC)

PI:   Stephen A Murray, PhD  NIH Report

The revolution in gene editing technology promises to transform the development of therapeutics to treat human disease. As part of the Somatic Cell Genome Editing consortium, the goal of this project is to build mouse resources and provide an animal model testing platform to support the optimization of novel genome editing technologies for future translational applications.
Summary of data submissions:
  • Data for 2 validation experiments were submitted on 2021-04-01 SCGE ID:1044
  • Data for 1 validation experiments were submitted on 2021-04-01 SCGE ID:1046
  • Data for 3 new model experiments were submitted on 2022-08-11 SCGE ID:1068
  • Data for 1 new model experiments were submitted on 2022-10-19 SCGE ID:1075
  • Data for 1 new model experiments were submitted on 2022-10-19 SCGE ID:1076
  • Data for 2 new model experiments were submitted on 2022-10-19 SCGE ID:1077
  • Data for 1 new model experiments were submitted on 2022-10-19 SCGE ID:1078
  • Data for 1 new model experiments were submitted on 2022-10-19 SCGE ID:1079
  • Data for 1 new model experiments were submitted on 2022-10-19 SCGE ID:1080

Submissions Details

VALIDATION - The Jackson Laboratory Gene Editing Testing Center - Chen Validation (Chen, Zheng-Yi)
SCGE ID:1044 - Submission Date: 2021-04-01 
Experiment Name Type Description
Independent validation of Chen delivery platform using LNPs to deliver CRISPR/Cas9 to mouse inner ear

In Vivo Delivery of CRISPR/Cas9 via bioreducible lipid nanoparticles (LNPs) to the inner ear in Ai14 mice
Repeat experiment of independent validation of Chen delivery platform using LNPs to deliver CRISPR/Cas9 to mouse inner ear

In Vivo Delivery of CRISPR/Cas9 editor via bioreducible lipid nanoparticle to the inner ear in Ai14 mice
VALIDATION - The Jackson Laboratory Gene Editing Testing Center - Gong Validation (Gong, Shaoqin)
SCGE ID:1046 - Submission Date: 2021-04-01 
Experiment Name Type Description
Independent validation of Gong delivery platform using RNP-loaded nanocages to deliver CRISPR/Cas9 to mouse brain

 In Vivo Delivery of CRISPR/Cas9 via RNP-loaded nanocages to the brain in Ai14 mice
NEW MODEL DEVELOPMENT - The Jackson Laboratory Gene Editing Testing Center (JAX-GETC)
SCGE ID:1068 - Submission Date: 2022-08-11 
Experiment Name Type Description
Validating Traffic Light Reporter 2 (TLR2) Mouse Model in Homozygous Blastocysts using SpCas9

 In Vitro Male and female homozygous mice mouse containing the Traffic Light Reporter 2 (TLR2) reporter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Cas9 RNP along with test guides and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for tagRFP fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 156-193. Total blast analyzed by Sanger sequencing, 84-120.
Validating Traffic Light Reporter 2 (TLR2) Mouse Model in Heterozygous Blastocysts using SaCas9

 In Vitro WT female mouse and homozygous male mouse containing the Traffic Light Reporter 2 (TLR2) reporter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Cas9 RNP along with test guides and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for tagRFP fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced. ICE analysis tool from Synthego was used to score edits. NOTE: Total blasts analyzed expressing the fluorescent reporter, 55-67. Total blast analyzed by Sanger sequencing, 55-67.
Validating Traffic Light Reporter 2 (TLR2) Mouse Model in Heterozygous Blastocysts using SpCas9

 In Vitro WT female mouse and homozygous male mouse containing the Traffic Light Reporter 2 (TLR2) reporter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Cas9 RNP along with test guides and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for tagRFP fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 46-120. Total blast analyzed by Sanger sequencing, 8-64.
NEW MODEL DEVELOPMENT - The Jackson Laboratory Gene Editing Testing Center (JAX-GETC) - GER10 small animal reporter, submission 1
SCGE ID:1075 - Submission Date: 2022-10-19 
Experiment Name Type Description
Validating Gene Editing Reporter 10 (GER10) Mouse Model in Heterozygous Blastocysts

 In Vitro WT female mouse and homozygous male mouse containing the Gene Editing Reporter 10 (GER10) reproter driven by the CAG promoter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Adenine Base Editor RNP complex and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for Venus (GFP) fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced to determine efficiency of base changes at the target region. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 174. Total blast analyzed by Sanger sequencing, 112.
NEW MODEL DEVELOPMENT - The Jackson Laboratory Gene Editing Testing Center (JAX-GETC) - GER12 small animal reporter, submission 1
SCGE ID:1076 - Submission Date: 2022-10-19 
Experiment Name Type Description
Validating Gene Editing Reporter 12 (GER12) Mouse Model in Heterozygous Blastocysts

 In Vitro WT female mouse and homozygous male mouse containing the Gene Editing Reporter 12 (GER12) reporter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Cas9 RNP along with test guides and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for Katushka2S (RFP) fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 58-69. Total blast analyzed by Sanger sequencing, 51-52.
NEW MODEL DEVELOPMENT - The Jackson Laboratory Gene Editing Testing Center (JAX-GETC) - TLR7 small animal reporter, submission 1
SCGE ID:1077 - Submission Date: 2022-10-19 
Experiment Name Type Description
Validating Traffic Light Reporter 7 (TLR7) Mouse Model for NHEJ in Heterozygous Blastocysts

 In Vitro WT female mouse and homozygous male mouse containing the Traffic Light Reporter 7 (TLR7) reporter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Cas9 RNP along with test guides and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for Katushka2S (RFP) fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 169. Total blast analyzed by Sanger sequencing, 117.
Validating Traffic Light Reporter 7 (TLR7) Mouse Model for HDR in Heterozygous Blastocysts

 In Vitro WT female mouse and homozygous male mouse containing the Traffic Light Reporter 7 (TLR7) reporter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Cas9 RNP along with test guides and HDR donor template, and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for Katushka2S (RFP) fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 103. Total blast analyzed by Sanger sequencing, 64.
NEW MODEL DEVELOPMENT - The Jackson Laboratory Gene Editing Testing Center (JAX-GETC) - GER14 small animal reporter, submission 1
SCGE ID:1078 - Submission Date: 2022-10-19 
Experiment Name Type Description
Validating Gene Editing Reporter 14 (GER14) Mouse Model in Heterozygous Blastocysts

 In Vitro WT female mouse and homozygous male mouse containing the Gene Editing Reporter 14 (GER14) reporter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with various CRISPR/Cas9 combinations (or Cre) and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for Katushka2S (RFP) fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 46-164. Total blast analyzed by Sanger sequencing, 39-142.
NEW MODEL DEVELOPMENT - The Jackson Laboratory Gene Editing Testing Center (JAX-GETC) - GER18 small animal reporter, submission 1
SCGE ID:1079 - Submission Date: 2022-10-19 
Experiment Name Type Description
Validating Gene Editing Reporter 18 (GER18) Mouse Model in Heterozygous Blastocysts

 In Vitro WT female mouse and homozygous male mouse containing the Gene Editing Reporter 18 (GER18) reporter driven by the EF-1 alpha promoter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Cas9 RNP along with two guides and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for GFP fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced to determine efficiency of deleting the IVS2-654 splicing variant. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 61. Total blast analyzed by Sanger sequencing, 54.
NEW MODEL DEVELOPMENT - The Jackson Laboratory Gene Editing Testing Center (JAX-GETC) - GER19 small animal reporter, submission 1
SCGE ID:1080 - Submission Date: 2022-10-19 
Experiment Name Type Description
Validating Gene Editing Reporter 19 (GER19) Mouse Model in Heterozygous Blastocysts

 In Vitro WT female mouse and homozygous male mouse containing the Gene Editing Reporter 19 (GER19) reporter driven by the CAG promoter at the Rosa26 locus were mated. Females were super-ovulated and zygotes harvested. Zyotes were electroporated with Cas9 RNP along with two guides and were cultured 96 hours to blastocyst stage. Blastocysts were imaged for GFP fluorescence and scored for fluorescence signal. Blastocysts were then PCR amplified for the reporter allele and Sanger sequenced to determine efficiency of deleting the IVS2-654 splicing variant. ICE analysis tool from Synthego was used to score edits. NOTE: Some blasts did not PCR amplify well and it could not be determined if they had edits. Total blasts analyzed expressing the fluorescent reporter, 53. Total blast analyzed by Sanger sequencing, 48.