STOCK Sema6a^tm2.1Alk/J

Stock number: 039872
Product name: Sema6A^null
Research areas: Developmental Biology Research, Neurobiology Research, Research Tools, Sensorineural Research

Description

The Sema6A^null strain has a floxed exon 3 removed by Cre-recombination in the sema domain, transmembrane domain (TM), and cytoplasmic domain, (semaphorin) 6A (Sema6A) gene. Sema6A is a transmembrane protein that has many implicated functions, including axon guidance, cell migration, cancer progression, and myelination. The line is useful for studies of Sema6A in neural development and axonal guidance, in addition to studies addressing development and function of multiple organ systems.

Detailed description

The Sema6A^null mice have exon 3 removed in the sema domain, transmembrane domain (TM), and cytoplasmic domain, (semaphorin) 6A (Sema6A) gene. The recombination between loxP sites introduces a frameshift and multiple premature stop codons, preventing the expression of Sema6A. The donating investigator reported that the HA-Sema6A is not produced in homozygous Sema6A^null mice indicating it is a null allele. The homozygous mice are viable but with reduced fertility and with retinal defects phenotype. The line is useful for studies of Sema6A in neural development and axonal guidance, in addition to studies addressing the development and function of multiple organ systems.

Of note, the donating investigator has made available mice with Sema6A^flox [3xHA::loxP-exon3-loxP; Stock No. 039871] and its germline Cre recombined derivative Sema6A^null [retaining 3xHA with exon3 deleted; Stock No. 039872].

Development

A targeting construct was designed to insert a 3x hemagglutinin (HA) tag downstream of the signal peptide in exon 2 at the N-terminus of the extracellular domain of Sema6A and loxP sites in introns flanking exon 3. The targeting construct was injected into (129S6 x C57BL/6J)F1-derived ES cells. The frt-flanked neo cassette was removed by transient FLP recombinase expression in the ES cells. Then, the ES clones with correct insertion were verified by sequencing and introduced into recipient mice blastocysts. The chimeric mice were bred with C57BL/6 mice. The resulting male Sema6A^flox conditional knock-out mice (Stock No. 039871) were bred with germline Sox2^Cre-recombinase female mice (Stock No. 008454). The offspring were then backcrossed to C57BL/6 wildtype mice for five generations to segregate Sox2^Cre away from the Sema6A^null allele and maintained in C57BL/6 genetic background by donating laboratory (see SNP note below). Upon arrival at The Jackson Laboratory, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6J (Stock No. 000664) oocytes.

Of note, the donating investigator has made available mice with Sema6A^flox [3xHA::loxP-exon3-loxP; Stock No. 039871] and its germline Cre recombined derivative Sema6A^null [retaining 3xHA with exon3 deleted; Stock No. 039872].

In 2024, 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 males sent to The Jackson Laboratory Repository. Three of the 43 markers throughout the genome were segregating with an unknown source. Also, one marker on Chromosome 3 was segregating with 129. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed genetic background.

Allele

Symbol: Sema6a^tm2.1Alk
Name: targeted mutation 2.1, Alex L Kolodkin
MGI accession ID: MGI:7661008
Generation method: Targeted
Attributes: Null/Knockout; Epitope tag
Marker symbol: Sema6a
Marker name: sema domain, transmembrane domain (TM), and cytoplasmic domain, (semaphorin) 6A
Marker synonyms: A730020P05Rik; HT018; sema; SEMA6A1; Sema6A-1; Semaq; VIa
Chromosome: 18
Strain of origin: (129S6/SvEvTac x C57BL/6J)F1
Molecular note: The targeting construct inserted an hemagglutinin (HA) tag is knocked in downstream of the signal peptide in exon 2 at the N-terminus of the extracellular domain and floxed exon 3. The frt-flanked neo cassette was removed by crossing the FLP deleter line. Cre-mediated recombination removed exon 3.