School of Biomedical Sciences, CUHK

The CUHK Transgenic Core Service Centre provides a range of services to investigators in the Chinese University of Hong Kong. The mission of the facility is to generate, breed and preserve genetically-modified mice. Initated and Coordinated by the School of Biomedical Sciences, the CUHK Transgenic Core Service. Centre was set up on 1st of December 2010. Specifically, the Transgenic Core Service provide expertize for production of genetically engineered mice by pronuclear injection of DNA constructs, embryonic stem cell and iPS cell injection into blastocyst. We will maintain an archive of frozen embryos and spermatozoa from different types of transgenic and knockout animals, produced at the site or imported from other Institutions. We will also provide technical knowhow and consultations on how to produce transgenic and knockout mice.

For consultation on DNA constructs and cell lines please contact Ms Li Pik Shan for details, on a first-come-first-serve basis. You can contact us below:

Contact person:

Site Manager: Ms. Li, Pik Shan, Tel: 39434224, Email: psli99@cuhk.edu.hk
Consultant: Professor Kenneth Lee, Tel: 39436785, Email: kaholee@cuhk.edu.hk

Location:

G05, Shanghai Fraternity Association Research Services Centre.

Services currently or will be available:

A. Transgenic Injections:

1. Pronuclei injection
2. ES cell / iPS cell injection

B. Maintenance of Cell Library:

1. Sperm Freezing
2. Embryo Freezing

C. Rederivation:

1. Embryo transfer, from frozen or fresh

Pronuclei injection

The injection of DNA constructs into the pronuclei of fertilized eggs is the most common method for making transgenic animals. Fertilized eggs are collected from superovulated females and injected by DNA. Then they are transferred to pseudopregnant recipients. The offsprings will be tailed and genotyped for positive.

Basic Service Provides:

1. 2 sessions of fertilized eggs from superovulated females (5x for F1CC or 10x for C57Bl6)
2. 2 sessions of DNA injection
3. Transfer of microinjected embryos into pseudopregnant recipients
4. Tail biopsy of offsprings.

Guarantee:

• At least 200 eggs will be injected, or
• 3 founders (with DNA integration, not guarantee for expressions.) produced, or 30 pups produced

Investigator must provide:

1. Brief description of the construct and the phenotype expected (size of construct, circuler / linerized, lethal or not, expected stage).
2. At least 20ul of >200ng/ul DNA in Tris buffer, 280/260nm > 1.6.
3. Genotype results of any tail biopsy

Turnaround Times

The minimum time required to produce transgenic founders is approximately 8-9 weeks, upon the Center receive the DNA construct. This includes 2 weeks for preparing the egg donors, 3 weeks gestation time, and 3-4 weeks from birth to weaning. The tail biopsy will be hand over to investigators for genotyping. The offspring will be held for 1 month, while waiting for the results of genotyping.

ES cell/ iPS cell injection

Gene targeting is normally performed in mouse embryonic stem (ES) cells, in vitro. Successfully targeted cells will then be injected into mouse blastocyst embryo to establish chimaeric mice. Chimaeric mice will be crossed with wild-type mice in order to elucidate whether the injected ES cells have integrated into the germline i.e. whether they can contribute to the germline of the chimaeric mice and produce an offspring from an ES cell-derived gamete.

Basic Service Provide:

1. 2 sessions of blastocyst harvest from superovulated females (10x for C57Bl6)
2. 2 sessions of ES/IPS cell injection
3. Transfer of injected blastocysts into pseudopregnant recipients

Guarantee:

• at least 50 blastocystsinjected, or
• 2 chimaeras or 30 pups produced

Investigator will provide:

1. Brief description of the targeted cell line and its genetic background
2. Trysinized targeted ES cells (without MEF ) in culture medium with 20mM HEPES on ice

Turnaround Times

The minimum time required to produce chimaera is approximately 8-9 weeks, upon receiving the cell line. This includes 2 weeks for preparing the egg donors, 3 weeks gestation time, and 3-4 weeks from birth to weaning.

Prices:

Transgenic Injection:

DNA Injection	2x injections (>200 oocytes) or 3 founders	HKD 10,000
	+ 2x injections (for serial embryo collection and analysis)	HKD 5,000
ES / iPS cell Injection	Blastocyst, 2x injection (>50 injected), or 3 chimeras	HKD 20,000

Cell Storage:

Sperm freezing	5 years per line	HKD 2,000
Embryo freezing	5 years per line	HKD 5,000

• Half number of frozen vials will be given back to investigator for back up keeping.