Recombinant constructs and transgenic fluorescent ornamental fish therefrom
Inventors
Blake, Alan • Crockett, Richard • Essner, Jeffrey • Hackett, Perry • Nasevicius, Aidas
Assignees
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Publication Number
US-9271478-B2
Publication Date
2016-03-01
Expiration Date
Abstract
The present invention relates to the method and use of reef coral fluorescent proteins in making transgenic red, green and yellow fluorescent zebrafish. Preferably, such fluorescent zebrafish are fertile and used to establish a population of transgenic zebrafish and to provide to the ornamental fish industry for the purpose of marketing. Thus, new varieties of ornamental fish of different fluorescence colors from a novel source are developed.
Core Innovation
The invention relates to producing a transgenic, fluorescent fish with improved fluorescence by generating chromosomally introduced transgenes carried on a single vector. The transgenic fish comprises a first transgene encoding a fluorescent protein under the control of a ubiquitous fish promoter and a second transgene encoding the same fluorescent protein under the control of a tissue-specific fish promoter.
The approach integrates both transgenes positioned on the single vector into the fish by chromosomal introduction, such that the fish exhibits fluorescence from both promoter contexts. The tissue-specific and ubiquitous expression modes are used together to achieve strong visible fluorescence distributed across the body.
The invention includes selecting transgenic fish exhibiting strong visible fluorescence over 75% to 100% of the body of the fish, optionally excluding fins and eyes. The document describes ornamental fluorescent fish, including reef coral fluorescent proteins such as DsRed2, ZsGreen1, and ZsYellow1 expressed in transgenic zebrafish, preferably Golden zebrafish, and reports specific transgenic integration events deposited in ECACC.
Claims Coverage
The independent claim clm-00001 defines a two-part producing method that includes generating chromosomally introduced transgenic fluorescent fish using a single vector carrying two promoter-controlled transgenes for the same fluorescent protein and selecting fish with strong visible whole-body fluorescence (75%–100%, optionally excluding fins and eyes).
Chromosomal introduction of two transgenes on a single vector
Transgenic, fluorescent fish are generated by chromosomal introduction of a first and second transgene positioned on a single vector.
Same fluorescent protein under ubiquitous and tissue-specific promoters
The transgenic fluorescent fish comprises a first transgene encoding a fluorescent protein under the control of a ubiquitous fish promoter and a second transgene encoding the same fluorescent protein under the control of a tissue-specific fish promoter.
Selection for strong visible fluorescence across the body
Selecting a transgenic fish exhibiting strong visible fluorescence over 75% to 100% of the body of the fish, optionally excluding fins and eyes.
Overall, the claim coverage is focused on producing chromosomally introduced transgenic fluorescent fish in which the same fluorescent protein is driven by both a ubiquitous fish promoter and a tissue-specific fish promoter from a single vector, followed by selecting fish that meet a strong visible fluorescence distribution criterion.
Stated Advantages
Improved fluorescence, including strong visible fluorescence across 75% to 100% of the body of the fish.
Documented Applications
Ornamental fluorescent fish for the ornamental fish industry, including transgenic zebrafish described as market/ornamental use cases.
Downstream distribution to the ornamental fish market is covered by dependent claim activity.
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