B6;129-Igk^{em1(IGK*)Abtx}/Igk^em2Abtx Igh^{em1(IGH*)Abtx}/Igh^em2Abtx/J

Stock number: 039317
Product name: Atlas™ Full Human Diversity Mouse
Strain synonyms: Fully-Hu KLC ; KLaP (k-KO) :: Fully human HC ; mIgDJ_D KO
Research areas: Cancer Research, Immunology, Inflammation and Autoimmunity Research, Research Tools

Description

The Atlas™ Full Human Diversity Mouse is a platform for generating chimeric monoclonal antibodies. When immunized, these mice produce antibodies with a human variable region and a mouse constant region.  The humanized variable regions are in both the heavy chain and the kappa light chain.  The strain includes >99% of commonly expressed human sequences in these variable regions.

Customers must have the appropriate agreement in place to purchase this strain. For more information, please contact Atlas@jax.org.

The Atlas™ Binary Fixed Light Chain Mouse for developing bispecific antibodies is also available under Stock No. 039319.

Detailed description

Atlas™ Full Human Diversity Mice contain targeted knock-ins of the human heavy and kappa light V(D)J regions in place of the endogenous variable region of the murine immunoglobulin loci.

By containing the near full complement of human antibody gene sequence diversity with antibody titers comparable to wildtype mice, Atlas™ Mouse antibodies are selected for and matured in vivo, simplifying downstream engineering and streamlining the discovery and development process. This unique platform exhibits the improved speed, diversity, developability, and affinity that are core to knock-in antibody mice.

Key Benefits:

• In vivo selection and maturation of antibodies resulting in higher numbers of developable lead antibody candidates with high affinity and fewer requirements for ex vivo engineering compared to in vitro display and wildtype mouse platforms.
• Variable regions comprising >99% of commonly expressed human sequences reduce the risk of immunogenicity and improve antibody developability characteristics by limiting the need for downstream engineering.
• Knock-in format into endogenous mouse loci allows for physiologically-relevant expression and generation of antibody sequences.
• Use of only functional variable heavy chain (VH) and kappa light chain (VK) gene segments, without pseudogenes or duplicates, promotes more productive B-cell immunoglobulin gene rearrangements.

Development

The Atlas™ Full Human Diversity Mouse (Stock No. 039317) is distributed as F1 (first filial generation) mice resulting from breeding the double homozygous Fully-Hu KLC :: Fully human HC strain (B6;129-Igk^{em1(IGK*)Abtx} Igh^{em1(IGH*)Abtx}/J) with the double homozygous KLaP (k-KO) :: mIgDJ_D KO strain (B6;129-Igk^em2Abtx Igh^em2Abtx/J). The resulting F1 mice are compound heterozygous at the Igk and Igh loci - each of the four mutations is described below.

The Fully-Hu KLC allele (Igk^{em1(IGK*)Abtx}) has the mouse immunoglobulin kappa chain complex (Igk) region variable (V) and joining (J) segments deleted and replaced with a human IGK V-J sequence. Specifically, CRISPR/Cas9 genome editing in 129S6/SvEvTac-derived CMTI-1 embryonic stem (ES) cells was used in two separate steps to deliver into the mouse Igk locus a ~108 kb sequence comprising human IGK V-J segments (a synthetic DNA construct containing an array of 34 IGK V segments plus all 5 IGK J segments). The transcript ID / genome build used as reference for the sequence coordinates or guide/donor sequences were not provided. The mouse Igk 3' enhancer and constant regions remain telomeric to the insertion. The allele also retains a hygromycin resistance cassette centromeric of the insertion. Sequence confirmed founders were bred to C57BL/6N mice for germline transmission, and later IVF-expanded using C57BL/6J female donors. The resulting mice were maintained on a mixed C57BL/6N;C57BL/6J;129S6 genetic background before being bred with the C57BL/6N;C57BL/6J;129S6 Fully human HC strain.
Of note, the hygromycin resistance cassette sequence present in the Fully-Hu KLC allele is unique and does not amplify using The Jackson Laboratory's current Generic Hygro assay primers [August 2024].

The Fully human HC allele (Igh^{em1(IGH*)Abtx}) has the mouse immunoglobulin heavy chain complex (Igh) region variable (V), diversity (D) and joining (J) segments deleted and replaced with a human IGH V-D-J sequence. Specifically, CRISPR/Cas9 genome editing in 129S6/SvEvTac-derived CMTI-1 embryonic stem (ES) cells was used in two separate steps to deliver into the mouse Igh locus a ~139 kb sequence containing an array of 41 IGH V segments plus a fully functional complement of 27 IGH D and 6 IGH J gene segments. The transcript ID / genome build used as reference for the sequence coordinates or guide/donor sequences were not provided. The mouse Adam6b/Adam6a loci remain telomeric to the human insertion, and the mouse Igh heavy chain constant regions (μ, δ, γ, ε, α) remain centromeric to the insertion. The allele also retains both puromycin resistance and hygromycin resistance cassettes telomeric of the insertion. Sequence confirmed founders were bred to C57BL/6N mice for germline transmission, and later IVF-expanded using C57BL/6J female donors. The resulting mice were maintained on a mixed C57BL/6N;C57BL/6J;129S6 genetic background before being bred with the C57BL/6N;C57BL/6J;129S6 Fully-Hu KLC strain or the C57BL/6N;C57BL/6J;129S6 Fixed LC strain.
Of note, the hygromycin resistance cassette sequence present in the Fully human HC allele is unique and does not amplify using The Jackson Laboratory's current Generic Hygro assay primers [August 2024].

The KLaP (k-KO) allele (Igk^em2Abtx) has the mouse immunoglobulin kappa chain complex (Igk) region variable (V) and joining (J) segments deleted. Specifically, CRISPR/Cas9 genome editing in 129S6/SvEvTac-derived CMTI-1 embryonic stem (ES) cells was used to remove a ~3.2 Mb sequence including the mouse Igk V and J segments and replace it with a PGK-neomycin resistance cassette. The transcript ID / genome build used as reference for the sequence coordinates or guide/donor sequences were not provided. The mouse Igk 3' enhancer and constant regions remain telomeric to the insertion. Sequence confirmed founders were bred to C57BL/6N mice for germline transmission, and later IVF-expanded using C57BL/6J female donors. The resulting mice were maintained on a mixed C57BL/6N;C57BL/6J;129S6 genetic background before being bred with the C57BL/6N;C57BL/6J;129S6 mIgDJ_D KO strain.

The mIgDJ_D KO allele (Igh^em2Abtx) has the mouse immunoglobulin heavy chain complex (Igh) region diversity (D) and joining (J) segments deleted. Specifically, CRISPR/Cas9 genome editing in 129S6/SvEvTac-derived CMTI-1 embryonic stem (ES) cells was used to remove a ~55 kb mouse Igh D-J segment and replace it with a PGK-neomycin resistance cassette. The transcript ID / genome build used as reference for the sequence coordinates or guide/donor sequences were not provided. The mouse Adam6b/Adam6a loci remain telomeric to the deleted region, and the mouse Igh heavy chain constant regions (μ, δ, γ, ε, α) remain centromeric to the deleted region. Sequence confirmed founders were bred to C57BL/6N mice for germline transmission, and later IVF-expanded using C57BL/6J female donors. The resulting mice were maintained on a mixed C57BL/6N;C57BL/6J;129S6 genetic background before being bred with the C57BL/6N;C57BL/6J;129S6 KLaP (k-KO) strain.

Allele

Symbol: Igk^{em1(IGK*)Abtx}
Name: endonuclease-mediated mutation 1, Dan Rohrer
MGI accession ID: MGI:7715231
Generation method: Endonuclease-mediated
Attributes: Inserted expressed sequence; Humanized sequence
Synonyms: Fully-Hu KLC
Marker symbol: Igk
Marker name: immunoglobulin kappa chain complex
Marker synonyms: kappa
Chromosome: 6
Strain of origin: 129S6/SvEvTac
Molecular note: The allele has the mouse immunoglobulin kappa chain complex (Igk) region variable (V) and joining (J) segments deleted and replaced with a human IGK V-J sequence. Specifically, CRISPR/Cas9 genome editing in 129S6/SvEvTac-derived CMTI-1 embryonic stem (ES) cells was used in two separate steps to deliver into the mouse Igk locus a ~108 kb sequence comprising human IGK V-J segments (a synthetic DNA construct containing an array of 34 IGK V segments plus all 5 IGK J segments).

Allele

Symbol: Igk^em2Abtx
Name: endonuclease-mediated mutation 2, Dan Rohrer
MGI accession ID: MGI:7715232
Generation method: Endonuclease-mediated
Attributes: Null/Knockout
Synonyms: KLaP (k-KO)
Marker symbol: Igk
Marker name: immunoglobulin kappa chain complex
Marker synonyms: kappa
Chromosome: 6
Strain of origin: 129S6/SvEvTac
Molecular note: The allele has the mouse immunoglobulin kappa chain complex (Igk) region variable (V) and joining (J) segments deleted. Specifically, CRISPR/Cas9 genome editing in 129S6/SvEvTac-derived CMTI-1 embryonic stem (ES) cells was used to remove a ~3.2 Mb sequence including the mouse Igk V and J segments and replace it with a PGK-neomycin resistance cassette.

Allele

Symbol: Igh^{em1(IGH*)Abtx}
Name: endonuclease-mediated mutation 1, Dan Rohrer
MGI accession ID: MGI:7715238
Generation method: Endonuclease-mediated
Attributes: Inserted expressed sequence; Humanized sequence
Synonyms: Fully human HC
Marker symbol: Igh
Marker name: immunoglobulin heavy chain complex
Chromosome: 12
Strain of origin: 129S6/SvEvTac
Molecular note: The allele has the mouse immunoglobulin heavy chain complex (Igh) region variable (V), diversity (D) and joining (J) segments deleted and replaced with a human IGH V-D-J sequence. Specifically, CRISPR/Cas9 genome editing in 129S6/SvEvTac-derived CMTI-1 embryonic stem (ES) cells was used in two separate steps to deliver into the mouse Igh locus a ~139 kb sequence containing an array of 41 IGH V segments plus a fully functional complement of 27 IGH D and 6 IGH J gene segments.

Allele

Symbol: Igh^em2Abtx
Name: endonuclease-mediated mutation 2, Dan Rohrer
MGI accession ID: MGI:7715239
Generation method: Endonuclease-mediated
Attributes: Null/Knockout
Synonyms: mIgDJ_D KO
Marker symbol: Igh
Marker name: immunoglobulin heavy chain complex
Chromosome: 12
Strain of origin: 129S6/SvEvTac
Molecular note: The allele has the mouse immunoglobulin heavy chain complex (Igh) region diversity (D) and joining (J) segments deleted. Specifically, CRISPR/Cas9 genome editing in 129S6/SvEvTac-derived CMTI-1 embryonic stem (ES) cells was used to remove a ~55 kb mouse Igh D-J segment and replace it with a PGK-neomycin resistance cassette.