The Vil-LacZ knock-in/knockout allele (also called Villinβ-gal) was designed to both abolish Vil1 gene function and express a fusion transcript encoding the 18 N-terminal amino acids of villin fused in-frame to the β-galactosidase cDNA. Expression of lacZ is directed to villus and crypt epithelial cells, along the entire small intestine and colon, in a cephalocaudal gradient similar to the expression pattern of endogenous Vil1. These mice may be useful in studying intestinal organogenesis, gastric stem cell biology and cancer.
Deborah L Gumucio, University of Michigan
Genetic Background | Generation |
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|
Allele Type | Gene Symbol | Gene Name |
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Targeted (Reporter, Null/Knockout) | Vil1 | villin 1 |
In adult mice, the villin 1 gene (Vil1) is highly expressed specifically in the intestinal epithelial cells, whereas the gastric epithelium is largely devoid of villin. The Vil-LacZ knock-in/knockout allele (also called Villinβ-gal) has lacZ replacing exons 1-3 of the Vil1 locus. This abolishes endogenous gene function and results in the villin 1 promoter/enhancer regions directing expression of a fusion transcript encoding the 18 N-terminal amino acids of villin fused in-frame to the β-galactosidase cDNA. No villin protein expression from this null allele is observed in intestinal epithelial tissue. Homozygous villin-deficient mice are viable and fertile with no gross abnormalities. Subtle ultrastructural alterations in the microvillar actin core of the intestine are reported in homozygotes. Villin/lacZ expressing cells are easily detectable by X-gal staining, both in whole embryos and in sections. Specifically, lacZ expression is primarily in villus and crypt epithelial cells, along the entire small intestine and colon, in a cephalocaudal gradient similar to the expression pattern of endogenous Vil1. Additionally, a rare population of gastric progenitor cells (GPCs) in the antrum are lacZ positive in an otherwise lacZ negative adult gastric epithelium.
The Vil-LacZ knock-in/knockout allele (also called Villinβ-gal) was designed by Dr. Deborah L. Gumucio (University of Michigan). A targeting vector was designed to replace exons 1-3 of the villin 1 locus (Vil1) with a cDNA encoding β-galactosidase (lacZ) and a reverse-oriented PGK-neomycin resistance cassette. The construct was electroporated into (129X1/SvJ x 129S1/Sv)F1-Kitl+)-derived R1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into recipient blastocysts, and chimeric males were bred with C57BL/6J females for germline transmission. The donating investigator reports that the mutant colony was maintained by breeding heterozygous males with C57BL/6NCrl females for more than ten generations (see SNP results below). In 2016, heterozygous males with black coat color were sent to The Jackson Laboratory Repository. Upon arrival, sperm was cryopreserved. To establish our live colony, an aliquot of frozen sperm was used to fertilize C57BL/6NJ oocytes (Stock No. 005304).
Of note, it is not known if the Y chromosome has been fixed to the C57BL/6 background during backcrossing.
In 2016, a 32 SNP (single nucleotide polymorphism) panel analysis, with 27 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 a cohort of the first generation rederived living colony at The Jackson Laboratory Repository. 26 of 27 markers throughout the genome suggested a C57BL/6 genetic background - the only exception was the marker in the panel closest to the Vil1 gene (segregating C57BL/6;129S allele-type). In addition, 4 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating in some mice (chromosomes 8 [~15.2 Mbp], 13 [~41.0 Mbp], 15 [~57.6 Mbp] and 19 [~49.9 Mbp]). These data suggest that the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6J;C57BL/6N genetic background.
Expressed Gene | lacZ, beta-galactosidase, E. coli |
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Site of Expression | Gastric progenitor cells. |
Allele Name | targeted mutation 1, Deborah L Gumucio |
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Allele Type | Targeted (Reporter, Null/Knockout) |
Allele Synonym(s) | |
Gene Symbol and Name | Vil1, villin 1 |
Gene Synonym(s) | |
Expressed Gene | lacZ, beta-galactosidase, E. coli |
Site of Expression | Gastric progenitor cells. |
Strain of Origin | (129X1/SvJ x 129S1/Sv)F1-Kitl+ |
Chromosome | 1 |
Molecular Note | A targeting construct containing lacZ and neo replaced 4.6 kb of genomic sequence encompassing a portion of the first coding exon (exon 1) and the following 2 exons (exons 2 and 3). An upstream non-translated exon was left intact. While low levels of aberrant transcipt were detected in mutant mice, neither normal nor truncated protein was detected by Western blot analysis. |
When maintaining a live colony, heterozygous mice may be bred together, to wildtype mice from the colony or to inbred mice (either C57BL/6J [Stock No. 000664] or C57BL/6NJ [Stock No. 005304]). Alternatively, homozygous mice may be bred together.
When using the Vil-lacZ knock-in/knock-out mouse strain in a publication, please cite the originating article(s) and include JAX stock #028946 in your Materials and Methods section.
Facility Barrier Level Descriptions
Service/Product | Description | Price |
---|---|---|
Herterozygous or wildtype for Vil1<tm1Gum> |
Frozen Mouse Embryo | B6.129-Vil1<tm1Gum>/J Frozen Embryo | $2595.00 |
Frozen Mouse Embryo | B6.129-Vil1<tm1Gum>/J Frozen Embryo | $2595.00 |
Frozen Mouse Embryo | B6.129-Vil1<tm1Gum>/J Frozen Embryo | $3373.50 |
Frozen Mouse Embryo | B6.129-Vil1<tm1Gum>/J Frozen Embryo | $3373.50 |
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