Transgenic and Knockout Mouse

Leadership

Cheryl Bock
Transgenic Facility Manager
919-684-3197
cbock@duke.edu

Meilang Flowers
Support Staff
Meiling.flowers@duke.edu

Gary Kucera, MS
BAC Recombineering Core Manager
gary.kucera@duke.edu

Traci Reddick, MS
Rederivation Core Manager
traci.reddick@duke.edu

Location

Genome Sciences Research Building 3022 and 3038

Resource Overview

The Transgenic and Knock Out Mouse Shared Resource provides complete services for the production of designer mutations in mice.

The resource specializes in microinjection of mouse embryos to create novel mutants and is staffed by experts in mouse embryonic stem cell culture.

Microinjection of DNA constructs into the pronucleus of fertilized mouse embryos produces founder mice typically known as “transgenic mice.” Microinjection of embryonic stem cells with altered genes into the mouse blastocyst produces mice that are colloquially known as “knockout mice.”

Using ES cell mediated mutagenesis, the resource has made mutant mice with point mutations, large deletions, conditional mutations, and gene exchanges (knock-ins) in addition to null mutations, or knockouts.

The resource is funded by the Duke Cancer Institute Core Grant and user fees.

Capabilities

• Microinjection of DNA into embryos
• Microinjection of ES cells into blastocysts
• Transfection and selection of ES cells
• PCR identification of targeted ES clones
• Tetraploid complementation

BAC Recombineering Core

The BAC Recombineering Core, a new branch of the resource, provides full service in design and production of DNA constructs using available Bacterial Artificial Chromosomes (BACs).

Recombineering (recombination-mediated genetic engineering) is a novel and powerful method for fast and efficient construction of vectors for subsequent manipulation of mouse genomes. It is based on homologous recombination in E. coli using recombination proteins provided from lambda phage.

This technical breakthrough is precise and independent of the presence of restriction sites and the size of the DNA molecule to be modified.

Recombineering in BACs is incredibly versatile and can be used to create vectors for gene knockout, conditional knockout, knockin, and modified BACs for transgenic mouse production. BAC constructs for production of transgenic zebrafish have also been produced.

This core is currently funded by a subsidy from the Transgenic Mouse Facility, user fees, and funds from Nancy Andrews, MD, PhD, dean of the School of Medicine, in support of the Cancer Institute mission.

Capabilities

• Design of targeting strategy
• Creation of DNA constructs using BACs
• Characterization of BAC clones via Pulse Field Gel Electrophoresis
• Design of PCR and Southern strategies for screening founder animals
• Knockouts, Knockins and Conditional Knockouts possible
• Constructs produced for creation of transgenic zebrafish or mice

Rodent Genetic Services Rederivation Core

The Rodent Genetic Services Rederivation Core is funded by the Division of Laboratory Animal Resources and the Duke Cancer Institute Core Grant, plus user fees.

Capabilities

• Cryopreservation of sperm or embryos
• in vitro fertilization
• Recovery of frozen embryos