The Thy1-Brainbow 1.1 transgene was designed with the mouse Thy1 regulatory elements surrounding four fluorescent proteins (orange (OFP), red (RFP), enhanced yellow (EYFP), and cyan (CFP)) sequences uniquely flanked with pairs of incompatible Lox sites. Specifically, this Brainbow 1.1 coding region contained (from 5’ to 3’) a LoxN site, Lox2272 site, LoxP site, Kusabira-Orange OFP (kOFP; with polyA sequence), LoxN site, mCherry RFP (with membrane tethering palmitoylation sequence and polyA sequence), Lox2272 site, monomeric EYFP (mYFP^A206K; with membrane tethering palmitoylation sequence and polyA sequence), LoxP site, mCerulean CFP (with membrane tethering palmitoylation sequence and 5’ FRT site and polyA sequence). This transgene was microinjected into C57BL/6JxCBA hybrid oocytes. Founder mice (line M) were bred with C57BL/6. The donating investigator reported that the resulting mutant mice were backcrossed to C57BL/6 (see SNP note below) for at least 5 generations prior to arrival at The Jackson Laboratory. Upon arrival, mice were bred to C57BL/6J (Stock No. 000664) for at least one generation to establish the colony.

Kusabira-Orange (KO or kOFP) is a true OFP with an artificial amino acid sequence attached to the N-terminus of the protein allowing bright orange fluorescence in cells. The mCherry RFP is an optimized DsRed/mRFP1 fluorescent protein variant with a combination of amino acid substitutions designed to improve spectral properties. The monomeric EYFP (mYFP^A206K) has an amino acid substitution replacing a hydrophobic region with a positively charged residue designed to prevent dimerization. The monomeric Cerulean (mCerulean) is a variant of ECFP (ECFP^{S72A/Y145A/H148D/A206K}) with amino acid substitutions designed to improve spectral properties and prevent dimerization.

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 the rederived living colony at The Jackson Laboratory Repository. While the 27 markers throughout the genome suggested a C57BL/6 genetic background, 4 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6J ; C57BL/6N genetic background.

In this line, animals from Stock No. 007911 were backcrossed to DBA/2J for at least 10 generations by the donating laboratory.

• Livet J; Weissman TA; Kang H; Draft RW; Lu J; Bennis RA; Sanes JR; Lichtman JW. 2007. Transgenic strategies for combinatorial expression of fluorescent proteins in the nervous system. Nature 450(7166):56-62PubMed: 17972876MGI: J:125961