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| Caption | (a-f) Schematic representation of the entire mouse Fras1 genomic region and targeting strategy used to produce the Fras1flox conditional allele, Fras1tm1.1Pjsc, and the Fras1delta2 allele, Fras1tm1.2Pjsc. (a) Fras1 gene locus encompassing 74 exons, with arrow head indicating the ATG start codon. A red star marks the 32 bp exon 2 to be targeted and a red rectangle indicates the region of the gene targeting expanded in b-f. (b) Wild-type allele and (c) targeting vector with 5' and 3' homology arms and the loxP arm flanking exon 2 indicated. One loxP site is inserted 5' of exon 2 and another 3' of the PGK-neo cassette (c). LoxP sites are denoted by triangles. PvuII, EcoRV, and ScaI restriction sites were introduced 5' and 3' of exon 2 to facilitate identification of wild-type versus recombined clones with relevant probes. (b, d-f) Schematic representation of Southern blot screening strategy. Wild-type (b) and targeted DNA fragments (d-e) are shown with the probes designated for screening. Double headed arrows indicate the size and position of the restriction fragments detected. Probe locations are indicated by the black rectangles. (d) The initial targeted allele. Probes were designed for identification of ESC clones harboring the 5' loxP site (PvuII and ScaI restriction sites for 5' and 3' HA probes respectively). An EcoRV site was introduced at the 5' end of the 3' homology arm for genomic screening. Wild-type and targeted fragments were detected with 5' (PvuII digest) and 3' (EcoRV digest) external probes detecting wild-type fragments of 9.8 kb and 15.5 kb, respectively, and 5.6 kb and 11.6 kb targeted fragments of Fras1flneo allele (b, d). The 3' HA probe (ScaI digest) facilitated detection of wild-type (10.5 kb fragment), floxed (7.2 kb fragment) and null Fras1 alleles (6.7 kb fragment) (b, e, f). Correct targeting of Fras1 mutated allele was additionally confirmed with a probe specific for the PGKneo cassette in conjunction with ScaI digest (8.9 kb fragment) (d). The size of the wild-type and mutant bands are indicated. | ||||||
| Copyright | This image is from Pitera JE, Genesis 2012 Dec;50(12):892-8, and is displayed with the permission of Wiley-Blackwell, who owns the Copyright. J:190659 | ||||||
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Associated Alleles |
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Mouse Genome Database (MGD), Gene Expression Database (GXD), Mouse Models of Human Cancer database (MMHCdb) (formerly Mouse Tumor Biology (MTB)), Gene Ontology (GO) |
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last database update 12/10/2024 MGI 6.24 |
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