The Rosa-CAG-LSL-Synaptophysin-tdTomato-WPRE targeting vector was designed with (from 5' to 3') a CMV-IE enhancer/chicken beta-actin/rabbit beta-globin hybrid promoter (CAG), an FRT site, a loxP-flanked STOP cassette (with stop codons in all 3 reading frames and a triple polyA signal), a Synaptophysin-tdTomato fusion protein sequence, a woodchuck hepatitis virus post-transcriptional regulatory element (WPRE; to enhance the mRNA transcript stability), a bovine growth hormone polyA signal, and an attB/attP-flanked PGK-FRT-Neo-polyA cassette.

To create the Synaptophysin-tdTomato fusion protein sequence, a mouse synaptophysin cDNA sequence (encoding a 308 amino acid transcript with all but the first six amino acids of the full-length protein) was fused in-frame via a six nucleotide linker to the amino terminus of the tdTomato sequence (a non-oligomerizing DsRed fluorescent protein variant with a 12 residue linker fusing two copies of the protein (tandem dimer)).

The entire Rosa-CAG-LSL-Synaptophysin-tdTomato-WPRE targeting vector was inserted between exons 1 and 2 of the Gt(ROSA)26Sor locus via electroporation of (129S6/SvEvTac x C57BL/6)F1-derived G4 embryonic stem (ES) cells. Correctly targeted ES cells (clone Ai34) were selected. Chimeric mice were bred to PhiC31-expressing mice (C57BL/6J congenic background; see Stock No. 007743) to remove the PGK-FRT-Neo-polyA cassette and replace it with the recombined attB/attP site (attL). The resulting Ai34D (or Ai34Δneo) mice were bred with C57BL/6J wildtype mice for at least two more generations (and the PhiC31 gene was removed) prior to sending to The Jackson Laboratory Repository. Upon arrival, Ai34D mice were bred with C57BL/6J inbred mice (Stock No. 000664) for at least one generations to establish the colony.

Approximate Controls

• 000664 C57BL/6J

Additional Information

• Considerations for Choosing Controls

• Zeng H. 2011. Direct Data Submission 2011/04/15 MGI Direct Data SubmissionMGI: J:170755