STOCK Or8a1b^{tm1.1(COP4*/EYFP)Tboz}/J

Stock number: 021206
Product name: M72-IRES-ChR2-YFP
Research areas: Neurobiology Research, Research Tools, Sensorineural Research

Description

These M72-IRES-ChR2-YFP knock-in mice express a ChR2(H134R)-EYFP fusion gene from the olfactory receptor family 8 subfamily A member 1B promoter Or8a1b (formerly Olfr160. As such, the ChR2(H134R)-EYFP fusion protein is expressed in olfactory sensory neurons (OSN) expressing this specific odorant receptor, M72. This targeted mutation strain may be useful for studying olfactory transduction, olfactory circuitry, and olfactory perception by light stimulation.

Detailed description

These M72-IRES-ChR2-YFP mice express a ChR2(H134R)-EYFP fusion gene from the from the olfactory receptor family 8 subfamily A member 1B locus, Or8a1b (formerly Olfr160). Mice that are homozygous for the targeted mutation are viable and fertile. Only olfactory sensory neurons (OSN) expressing this specific odorant receptor, M72, express ChR2(H134R)-EYFP, rendering them light sensitive. Sensory neurons that express the modified allele express both ChR2-YFP as well as the intact M72 receptor and project axons to a medial and a lateral glomerulus on the dorsal surface of each olfactory bulb. Light directed at the glomerulus can elicit neuronal activity and drive odor guided behavior. These mutant mice thus allow precise, timed delivery of sensory input to the olfactory system. Homozygotes are viable and fertile. M72-IRES-ChR2-YFP mice are similar to STOCK Omp^{tm1.1(COP4*/EYFP)Tboz}/J mice (Stock No. 014173), which express ChR2(H134R)-EYFP all mature olfactory sensory neurons.

The ChR2(H134R)-EYFP fusion protein is composed of a Chlamydomonas reinhardtii-derived channelrhodopsin-2 that harbors a gain-of-function H134R substitution fused in-frame with an enhanced yellow fluorescent protein. The ChR2(H134R) is designed to cause larger stationary photocurrents compared to ChR2.

The bacterial opsins are retinal-binding proteins that combine a light-sensitive domain with an ion channel or pump; providing light-dependent ion transport, membrane potential alteration, and sensory functions to bacteria. This ChR2(H134R) functions as a blue light-driven cation channel that depolarizes the cell and causes action potentials. As such, illuminating ChR2(H134R)-expressing cells with blue light (450-490 nm) leads to rapid and reversible photostimulation of action potential firing activity in these cells.

Development

A targeting vector was designed to insert an internal ribosome entry site (IRES), the ChR2(H134R)-EYFP fusion gene, and a self-excising neo cassette, ACNF, downstream of the olfactory receptor family 8 subfamily A member 1B gene (Or8a1b). This construct was electroporated into 129-derived embryonic stem (ES) cells and correctly targeted ES cells were aggregated with C57BL/6J morulae. Resulting blastocysts were implanted in pseudopregnant females. Chimeric male offspring were crossed to B6(Cg)-Tyr^c-2J/J females (Stock No. 000058) to generate F1 heterozygous mice. These mice were bred together to generate homozygotes. Upon arrival at The Jackson Laboratory Repository, the mice were crossed to C57BL/6J mice (Stock No. 000664) for at least one generation to establish the colony. The Tyr^c-2J allele was bred out of the colony.

Allele

Symbol: Or8a1b^{tm1.1(COP4*/EYFP)Tboz}
Name: targeted mutation 1.1, Thomas Bozza
MGI accession ID: MGI:5469474
Generation method: Targeted
Attributes: Reporter
Marker symbol: Or8a1b
Marker name: olfactory receptor family 8 subfamily A member 1B
Chromosome: 9
Strain of origin: 129
Expressed genes: YFP,Yellow Fluorescent Protein,jellyfish
Site of expression: The ChR2(H134R)-EYFP fusion protein is expressed only in olfactory sensory neurons that express the specific odorant receptor, M72.
Molecular note: A targeting vector was designed to insert an internal ribosome entry site (IRES), the ChR2(H134R)-EYFP fusion gene, and a self-excising neo cassette, ACNF, downstream of the olfactory receptor 160 gene. The neo cassette is excised. Only olfactory sensory neurons (OSN) expressing this specific odorant receptor, M72, express ChR2(H134R)-EYFP, rendering them light sensitive.