B6.Cg-Tg(K18-ACE2)2Prlmn Ifnar1^tm1.2Ees/J

Stock number: 035041
Product name: K18-hACE2 ; Ifnar1^-
Research areas: Cell Biology Research, Developmental Biology Research, Immunology, Inflammation and Autoimmunity Research, Internal/Organ Research, Research Tools, Virology Research

Description

These mice express human ACE2, the receptor used by the severe acute respiratory syndrome coronavirus (SARS-CoV) to gain entry to cells. The human keratin 18 (KRT18) promoter directs expression to epithelia, including airway epithelial cells where infections typically begin. Recent research indicates the K18-hACE2 transgene may be useful in studying antiviral therapies for the 2019 novel coronavirus (SARS-CoV-2) pathogenesis and the disease outbreak COVID-19.

This strain also contains an Ifnar1^- targeted mutation which may result in reduced immune response and increased susceptibility to SARS-CoV viral infection during clinical trials.

Please note, these mice should be handled in a manner consistent with CDC/ABSA/WHO guidelines for prevention of human infection with the SARS-CoV-2 virus. Proper PPE and handling methods should be used at all times when working with these mice. Additional important guidelines for using SARS-CoV-2 susceptible mouse lines.

Detailed description

Please see the K18-hACE2 datasheet (Stock No. 034860) for important safety/handling and any updated information on K18-hACE2 mice in SARS-CoV-2 research.

Stock No. 035041 K18-hACE2 ; Ifnar1^- is a multiple mutant strain assembled by breeding Stock Nos. 034860 and 028288. In an attempt to offer popular alleles together, mouse lines may be bred together for purpose of researchers to study and characterize. It should be noted that the phenotype of Stock No. 035041 could vary from that originally described for the individual mouse lines used to assemble it. We may modify the strain description if necessary as published results become available. Each mutant allele is briefly described below [September 2020].

K18-hACE2 transgenic mice express the receptor for severe acute respiratory syndrome (SARS), caused by a coronavirus (SARS-CoV) in airway and other epithelia under control of the human keratin 18 (KRT18) promoter. Specifically, these mice contain 2.5 kb of the KRT18 genomic sequence, including the promoter, and the first intron (with a mutation in the 3' splice acceptor site to reduce exon skipping) and a translational enhancer (TE) sequence from alfalfa mosaic virus, upstream of the human angiotensin I converting enzyme (peptidyl-dipeptidase A) 2 coding sequence (hACE2), followed by exons 6-7 and the poly(A) signal of the human K18 gene. These elements have been found to be necessary for high-level expression and epithelial cell specificity of hACE2, the primary host cell receptor for SARS-CoV. In these mice, from founder line 2, K18-hACE2 transgene expression is evident in airway epithelial cells (but not in alveolar epithelia), as well as in epithelia of other internal organs, including the liver, kidney, and gastrointestinal tract. Recent research indicates that this line may also be useful in studies related to the study of 2019 novel coronavirus (SARS-CoV-2) pathogenesis.

Addition of the Ifnar1^- targeted mutation may result in reduced immune response and increased susceptibility to SARS-CoV viral infection during clinical trials.

The K18-ACE2 transgene and Ifnar1^- targeted mutation are able to be maintained as homozygous in their individual mouse lines. Stock No. 035041 is viable and fertile as hemizygous K18-ACE2 and heterozygous Ifnar1^-. However, it is not yet known if Stock No. 035041 may be maintained as homozygous for both alleles [September 2020].
Of note, if homozygous null Ifnar1^- animals are maintained in low health barrier rooms, the use of medicated water (e.g., sulfatrim/trimethoprim-sulfa or enrofloxacin/Baytril) is suggested to increase overall colony health.

Development

The ~6.8 kb K18-hACE2 transgene was designed with, from 5' to 3', 2.5 kb of genomic sequence, including the promoter, and the first intron (with a mutation in the 3' splice acceptor site to reduce exon skipping) from the human keratin 18 (KRT18) gene, and a translational enhancer (TE) sequence from alfalfa mosaic virus. This was followed by the human angiotensin I converting enzyme (peptidyl-dipeptidase A) 2 (hACE2) coding sequence, and exons 6-7 and the poly(A) signal of the human K18 gene. This transgene was microinjected into fertilized (C57BL/6J x SJL/J)F2 oocytes. K18-hACE2 mice from founder line 2, with 8 copies of the transgene, were subsequently maintained by breeding transgenic mice with C57BL/6 mice for at least two generations (the genetic background of the mice sent to The Jackson Laboratory was later determined to be C57BL/6N via SNP testing). Upon arrival, three backcrosses to C57BL/6J (Stock No. 000664) were then performed to rederive, establish and expand a congenic colony at The Jackson Laboratory as Stock No. 034860. Some of these mice were bred to Ifnar1^- mice (Stock No. 028288), carrying a knock-out mutation for the mouse Ifnar1 (interferon (alpha and beta) receptor 1) gene, to create this new strain as Stock No. 035041.

In May 2020, analysis commissioned by The Jackson Laboratory determined that multiple copies of the Tg(K18-ACE2)2Prlmn transgene integrated at a single site on chromosome 2 (99,209,508-99,220,724; mouse mm10). An ~11 kb duplication of the host genome (chr2:99,209,508-99,220,724) is present at both ends of the integrated transgene array. Currently, there are no genes annotated in this region. The transgene copy number of the hemizygous mouse is estimated to be 8 full copies (or 12-30 partial copies).

Allele

Symbol: Ifnar1^tm1.2Ees
Name: targeted mutation 1.2, Edward E Schmidt
MGI accession ID: MGI:5692167
Generation method: Targeted
Attributes: Null/Knockout
Marker symbol: Ifnar1
Marker name: interferon (alpha and beta) receptor 1
Marker synonyms: AVP; Ifar; IFN-alpha/betaR; IFN-alpha-REC; IFNAR; IFNBR; IFRC; IMD106
Chromosome: 16
Strain of origin: (C57BL/6 x 129S6/SvEvTac)F1
Molecular note: The targeting vector was designed to insert a self-excising FRT-Prm1pFlppol2Neo-FRT cassette and loxP site in intron 2, and a second loxP site in intron 3 of the gene. Facilitated self-excision of the cassette in the male germline and resulted in offspring with one FRT site and loxP site upstream of exon 3, and a second loxP site downstream of exon 3. Cre-mediated recombination removed the floxed neo cassette and exon 3. The resulting non-functional protein contains an exon 2-4 splice variant and frameshift leading to translation of 11 arbitrary (missense) amino acids within the N-terminal extracellular domain followed by a stop codon.

Allele

Symbol: Tg(K18-ACE2)2Prlmn
Name: transgene insertion 2, Stanley Perlman
MGI accession ID: MGI:6389235
Generation method: Transgenic
Attributes: Inserted expressed sequence; Humanized sequence
Synonyms: K18-hACE2
Marker symbol: Tg(K18-ACE2)2Prlmn
Marker name: transgene insertion 2, Stanley Perlman
Chromosome: 2
Strain of origin: (C57BL/6J x SJL/J)F2
Site of expression: Epithelia, including airway epithelia.
Molecular note: The ~6.8 kb transgene was designed with, from 5' to 3', 2.5 kb of genomic sequence, including the promoter, and the first intron (with a mutation in the 3' splice acceptor site to reduce exon skipping) from the human K18 gene, and a translational enhancer (TE) sequence from alfalfa mosaic virus. This was followed by the human angiotensin I converting enzyme (peptidyl-dipeptidase A) 2 (hACE2) coding sequence (lacking the last exon), and exons 6-7 and the poly(A) signal of the human K18 gene. The transgene integrated at a single site on chromosome 2 (99,209,508-99,220,724; mouse mm10). An ~11 kb duplication of the host genome (chr2:99,209,508-99,220,724) is present at both ends of the integrated transgene array. Currently, there are no genes annotated in this region. The transgene copy number of the hemizygous mouse is estimated to be 8 full copies (or 12-30 partial copies). Transgene expression is evident in airway epithelial cells (but not in alveolar epithelia), as well as in epithelia of other internal organs, including the liver, kidney, and gastrointestinal tract.