STOCK Lcn2^tm2Bara Tyr^c-2J/J

Stock number: 039157
Product name: Ngal-Luc2/mC
Research areas: Dermatology Research, Immunology, Inflammation and Autoimmunity Research, Metabolism Research, Mouse/Human Gene Homologs, Research Tools, Sensorineural Research

Description

Ngal-Luc2/mC knock-in/knock-out mice express a double-fusion reporter gene encoding luciferase-2 and mCherry (Luc2-mC) under the direction of the endogenous Lcn2 promoter. This mutation abolishes endogenous Lcn2 expression, while allowing for in vitro and in vivo bioluminescence/fluorescence imaging of NGAL expression. These mice may be useful for evaluating NGAL activity, in real-time, when studying pathogenesis of multiple inflammatory diseases, such as acute and chronic kidney disease.

Detailed description

Neutrophil gelatinase-associated lipocalin (NGAL), also referred to as Lipocalin-2 (LCN-2), is an iron-trafficking protein that is secreted mostly by immune cells such as neutrophils, macrophages, and dendritic cells. NGAL production is stimulated in response to inflammation, bacterial growth and infection. NGAL can be measured in plasma, urine, and other biological fluids and may be useful as a biomarker for kidney disease, peritonitis, atherosclerosis, hypertension, myocardial infarction, heart failure and some cancers.

These Ngal-Luc2/mC knock-in/knock-out (KI/KO) mice harbor a double-fusion reporter gene encoding luciferase-2 and mCherry (Luc2-mC) inserted upstream of the Lcn2 start codon. This mutation results in loss of endogenous NGAL production, while allowing for expression of Luc2-mC reporter proteins under the direction of the endogenous Lcn2 promoter.

Heterozygous and homozygous mice are viable and fertile and should be housed in a specific pathogen free (SPF) facility. Using this Ngal-Luc2/mC KI/KO mouse model, the donating laboratory was able to demonstrate cell-specific NGAL protein expression in response to unilateral kidney ischemia, in real time. Their data revealed that Ngal-Luc2-mC expression level was proportional to the severity of the injury and was detected as early as 3-6 hours post-injury via in vivo bioluminescence detection, which is significantly earlier than serum creatinine level changes after ischemic injury. These Ngal-Luc2/mC KI/KO mice may be useful for evaluating the expression pattern of NGAL in vivo when studying pathogenesis and therapeutic response in multiple inflammatory diseases, such as chronic kidney disease and cardiac myopathies.

Development

The Ngal-Luc2/mC knock-in/knock-out (KI/KO) allele was generated by insertion of a double-fusion reporter gene encoding luciferase-2 and mCherry (Luc2-mC) between the lipocalin 2 (Lcn2) start codon and exon 1. The targeting construct also included a loxP-flanked PGK-neomycin cassette. The construct was electroporated into (C57BL/6J x 129S6/SvEvTac)F1-derived KV1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts. The resulting chimeric mice were bred with B6(Cg)-Tyr^c-2J/J (B6-albino, Stock No. 000058) to establish germline transmission. The resulting heterozygous Ngal-Luc2/mC KI/KO mice, with floxed neo cassette intact, were maintained on a B6-albino background by the donating laboratory (see SNP note below). Upon arrival at The Jackson Laboratory, an aliquot of frozen sperm was used to fertilize B6(Cg)-Tyr^c-2J/J (B6-albino) oocytes.

in 2025, a 48 SNP (single nucleotide polymorphism) panel analysis, with 43 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the rederived living colony at The Jackson Laboratory Repository. Four of the 43 markers throughout the genome were segregating with 129. Also, the rd8 recessive mutation associated with retinal degeneration on Chromosome 1 within the Crb1 locus found to be segregating with C57BL/6N. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed genetic background.

Allele

Symbol: Lcn2^tm2Bara
Name: targeted mutation 2, Jonathan Barasch
MGI accession ID: MGI:5703928
Generation method: Targeted
Attributes: Reporter
Synonyms: Ngal-Luc2/mC
Marker symbol: Lcn2
Marker name: lipocalin 2
Marker synonyms: 24p3; MSFI; neu-related lipocalin; NGAL; NRL; p25; Sip24
Chromosome: 2
Strain of origin: (C57BL/6J x 129S6/SvEvTac)F1
Molecular note: The targeting construct introduced into a site between the 5'UTR and the start codon in exon 1 a DiFusion reporter consisting of a luciferase2 gene and mCherry followed by a floxed neomycin resistance cassette.

Allele

Symbol: Tyr^c-2J
Name: albino 2 Jackson
MGI accession ID: MGI:1855985
Generation method: Spontaneous
Attributes: Null/Knockout
Marker symbol: Tyr
Marker name: tyrosinase
Marker synonyms: ATN; CMM8; Oca1; OCA1A; OCAIA; SHEP3; skc35
Chromosome: 7
Strain of origin: B6.Cg-Tyrp1^b Hps1^ep
Site of expression: Expressed in melanosomes
Molecular note: This mutation has a G-to-T base change at coding nucleotide 230 resulting in an amino acid change from arginine to leucine at residue 77 (p.R77L) which lies in the highly conserved DDRE sequence. Nucleotide 230 is at the alternative 5' splice donor site for exon 1 and this allele does not produce the 1a or 1b subset of tyrosinase transcripts but does produce a significant increase in 1c and 1d transcripts. Western blots of homozygous mutant skin extracts demostrate the nearly complete absence of the broad 76-84 kDa band of glycosylated wild-type tyrosinase. No tyrosinase activity was found in hairbulb extracts from homozygous mice.