ºù«Ӱҵ

The zebrafish owes much of this success to its hardiness, cold resistance, easiness to breed and variable omnivorous diet.

This popular fish is well established as a powerful tool in the research of cellular and molecular mechanisms that underlie development and disease.

Genetic engineering is widely used, with much success, to recapitulate and track pathological processes.

The facility provides support for research with adequate provision of microscopes (both light and fluorescent), injection rigs, a needle puller, incubators (including light cycle incubators), behavioural analysis and all aquarium associated equipment.

Some tanks are equipped with light boxes so that day and night photoperiods can be changed, and some tanks are designed as coordinates for a multiwell plate. The full list of facilities includes:

• Behavioural Analysis Booths Suite
• Microinjection Rooms
• Microscope Rooms
• Programmable light boxes and light cycle incubators
• Quarantine Facilities
• Live food rearing provisions
• Cryogenic storage for IVF service

There are two quarantine facilities to support fish imports and a cold room to store all our feeds.

On this page

Diet | Stocks | Transgenics | Genotype servicing | Legislation | Regulation

---

Diet

Zebrafish are fed a varied diets that provides complete nutrition for all life stages. The fish receive a combination of specifically formulated dry feeds and live feeds.

The diet is aimed at maintaining excellent fish health, improving growth and preserving good reproductive performance. The swimming behaviour of the live feeds also provides environmental enrichment by encouraging the natural feeding behaviour of the fish.

Fry raising

Zebrafish fry are capable of independent feeding at five days post fertilisation (DPF), if kept in water at 28C. At this stage the fry are kept in 3.5L tanks of system water and fed the appropriate feed, as the fry grow and become capable of eating larger food particle sizes their diet is adjusted.

• 5-10 DPF rotifers and Gemma micro 75
• 11-31 DPF Artemia and Juvenile powder mix 75-200
• 32+ DPF Artemia, Gemma micro 300, Zebrafeed 400-600

---

Stocks

The Biological Services Aquarium (BSA) holds over 750 different zebrafish families called lines. Six of these lines are wild-type strains, which are similar to the Zebra Danio fish that you can buy in most tropical fish shops. 

The remaining lines are all genetically modified by genetic engineering to incorporate a transgene or genetic alteration in their genome. These lines enable advanced biological analysis of life course biology.

We keep stringent records of all fish in the facility including the date of birth, numbers in each tank and the tank location of where they are kept. We use a database to hold these records that is updated regularly with each change that happens.

Here at ºù«Ӱҵ the aquarium team not only perform the daily husbandry for the fish but we also maintain the majority of the lines we hold. We identify heterozygous fish by phenotype (their physical appearance) and also provide a genotyping service (to view their genetic appearance).

Zebrafish are past their optimum breeding age by the time they are two years old and so to maintain the line we breed new generations at around 14 months of age. They are usually fully mature and breeding well by the age of six months so this ensures that we always have a healthy breeding stock.

The majority of genetically modified fish are kept in heterozygous form (asymptomatic) and are crossed to wild-type to maintain good genetic variation. As only 50% of the offspring will carry the genetic change we often need to in-cross the fish and examine the offspring for the required relevant phenotype. Some of these lines do not produce a phenotype and so will require a small amount of the caudal fin to be taken and the DNA sent off for analysis, this is called genotyping.

Transgenic fish are more often kept in homozygous form (also asymptomatic) and are in-crossed to maintain the line, to stop these fish becoming unhealthy due to inbreeding they are also out-crossed to wild-type fish every two or generations. 

Transgenic fish can usually be selected before they reach the age of protection, at 5 days past fertilisation, and so do not need to be identified at a later stage.

We breed fish in two different ways

One method is to put a large box with a mesh insert into a shoaling tank of more than three fish. On top of the mesh we place marbles, as these resemble small pebbles that are often found in the natural habitat of the zebrafish. 

The lights coming on in a morning triggers the fish to begin breeding, the females will lay her eggs onto the marbles and the males will then swim over and fertilise them. Once the eggs have fallen through the mesh that adult fish cannot reach them, as there is a tendency for the eggs to be eaten. We can then take the spawning box out of the tank, remove the mesh and marbles and collect the eggs.

The other method is useful if you want to know who the exact parents are or you want the fish to breed at a particular time. A male and female are placed into a breeding box that has a grid at the bottom. This grid acts both to separate the adults from the eggs and also to resemble the natural habitat. A divider is placed between the two fish, this acts not only to stop any fighting that may occur overnight but also means that the fish will not breed until it is removed, although they can sense each other through the transparent divider. Once the divider is removed, after the lights have come on, breeding will commence.

---

Transgenics and the use of wild-types

We have approximately 400 transgenic fish lines within the aquarium.

A transgene is a gene or genetic material that has been transferred naturally, or by any of a number of genetic engineering techniques from one organism to another.

Transgenes are usually used to visualise a biological structure or process within an organism that can be studied in vivo. Fluorescent proteins are particularly useful. Green-fluorescent protein, for instance, is a jellyfish protein that has been extensively used to highlight the protein of interest in the biological process being studied.

The use of certain wild-type lines helps researchers to visualise transgenes in vivo. Most wild-type strains are derived from zebrafish that can be purchased in pet shops. There are Nacre, Casper, Long-tailed (TL)  as well as the ‘standard’ zebrafish termed e.g AB and London wild-type (LWT), which have the classic stripes.

The final wild-type lines have been recently derived from wild caught fish, such as WIK. The lack of pigmentation in the Nacre and Casper strains enable fluorescent proteins to be seen more clearly under a microscope. Long tailed strains have longer fins easier to fin clip, which may be taken for genotyping purposes.

Certain transgenic lines that express fluorescent proteins are visible in embryos between one and five days old under a microscope. These fluorescent proteins can be used to track changes in developmental processes and different coloured fluorescent proteins can be used in the same fish to further illustrate cellular processes in vivo.

---

Genotyping service 

The Biological Services Aquarium offers a genotyping service for all facility users. The service includes fin clipping, DNA extraction, PCR, restriction enzyme digest (or sequencing via RHH if required), interpretation of data results from agarose gels and relocation of identified fish ready for use for breeding.

We have specialised tanks with a genotyping rack for this service, providing water flow and feeding capability whilst housing up to 288 fish. After fin clipping, the turnaround time is typically the following day for results. The charge is based on consumable recover costs only and will free up researcher’s time utilizing this service.

The service is carried out by Lisa Van Hateren (email: l.j.gleadall@sheffield.ac.uk). Please email for request forms or for further information.

---

Legislation

The UK Home Office regulates the breeding and use of zebrafish at the BSA by granting an establishment licence under which named persons, such as the Veterinary Surgeon and Named Animal Care and Welfare Officers (NACWOs), can work.

Tight regulations and careful consideration of project proposals ensure only well-considered, ethically reviewed research is undertaken.

Home Office

The Animals (Scientific Procedures) Act 1986 Amendment Regulations 2012 (SI 2012/3039) amend the Animals (Scientific Procedures) Act 1986 (ASPA) to transpose European Directive 2010/63/EU on the protection of animals used for scientific purposes. The Directive sets out revised measures for the protection of animals used for scientific purposes.

This guidance is issued under the terms of section 21(1) of ASPA and provides information about the way in which the Secretaries of State for the Home Department and for Northern Ireland propose to exercise their powers under the Act. This guidance is for everyone involved with animals that are bred for, supplied for, or used in scientific procedures.

---

Regulation

Cefas

The Centre for Environment, Fisheries and Aquaculture Science - makes an important contribution to securing healthy and sustainable marine and freshwater environments so that current and future generations can prosper.

Yorkshire Water

Environmental officers advise us what we can and cannot put down the drain.

University of ºù«Ӱҵ

We abide by the local rules set out by the University of ºù«Ӱҵ these include employment right, health and safety, out-of-hours rules and fire training

Understanding Animals in Research

As a University we have signed thes