Transgenic Lines

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Most of the drawbacks of viruses can be overcome by permanent gene transfer via germline transgenesis. This approach aims at establishing lines of transgenic animals expressing the protein of interest stably and constitutively, eliminating the need of delivering the gene-of-interest on a single animal basis. When compared to viral approaches, the main issue of “optogenetic” transgenic lines so far has been their lower expression levels (notable exceptions are mouse lines using the thymus cell antigen 1 - Thy1 - promoter region and the Vglut2-ChR2-EYFP BAC transgenic line; see Hägglund et al., 2010). This limitation can be problematic for optogenetic tools requiring high expression levels such as microbial opsins. Nevertheless, a number of mouse strains expressing optical reporters (GCaMP, VSFPs, synaptopHluorin) or control tools (ChR2, VChR1, NpHR, eNpHR3) have been generated and the catalog of "optogenetic" mouse lines is expanding quickly. These lines express the optogenetic tool under the control of either a specific promoter (ChAT, VGAT, TPH2, VGluT2) or a Cre-activated cassette (e.g. Ai26 and Ai32 lines). The latter type of strain can be crossed with any existing Cre driver line to achieve targeted expression through cell-type specific recombination.

Although transgenesis clearly saves time and money on the long run, its implementation can be costly and time consuming. While this is true for classical transgenesis techniques (pronuclear microinjection and ES integration into blastocysts), new techniques such as testis electroporation Dhup and Majumbar, 2008, lentivirus-mediated and zinc finger nucleases-mediated transgenesis [1] might hold the keys for rapid, efficient and cost-effective germline transgenesis in various mammalian species.

Optogenetic control of locomotor activity in the Thy-1::ChR2-EYFP mouse

Classical Transgenesis

Microinjection of DNA into the pronucleus

Injection of ES cells into blastocyst-stage embryos

Transgenesis via electroporation of the testis

A paper by Dhup and Majumbar, 2008 describes an original technique for quick and cost-effective transgenesis using gene transfection in undifferentiated spermatogonia.

Commercial Suppliers of Transgenic rats

Genoway

SAGE Labs Rats

Updated Nov. 2011: SAGE Labs (Sigma Advanced Genetic Engineering Labs) are starting to produce knock-in and knock-out transgenic rats (mostly Sprague Dawley), including 5 CRE lines (Tryptophan hydroxylase, Tyrosine hydroxylase, Dopamine transporter, Parvalbumin and CaMKIIa). Price and availability are not known yet, but these rats look to be on target to be available at the end of 2012 for around $500 per animal.

Optogenetic Transgenic Mice from the Jackson Lab

References

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  1. Error fetching PMID 20015561: [LeProvost2010]