Targeted Transgenesis Services

Introduction

The standard Targeted Transgenesis service includes the following:
Linearization and purification of the targeting construct DNA.
Electroporation into ES cells.
Growth of cells under selection.
Picking at least 24 drug-resistant colonies into a 96-well plate.
Splitting the colonies into duplicate plates.
Freezing one plate and extracting DNA from the other plate.
Digestion of ES cell colony DNA with restriction enzyme.
Agarose gel electrophoresis and gel photography.
Capillary transfer of digested DNAs to nylon membrane.
Hybridization of membrane with radioactive probe (standard ROSA26 probe supplied by the TMF).
Washing blot and imaging on phosphorimager.
Analysis and annotation of image.

Turnaround Times

The minimum time needed to go from electroporation of the targeting construct to clone DNA ready for genotyping is about 4 weeks. The actual time may be somewhat longer if other targeting jobs are already in progress.
Southern blotting turnaround times can vary, depending on how many projects are in progress. At present, the minumum time is about 10 days.
Clone expansion and DNA extraction require an additional 2 weeks.

Advantages

This service offers the following advantages over the traditional method of pronuclear injection:
Similar levels of expression in all founder lines.
A known locus of integration.
Transgenes are always inserted as a single copy, minimizing the chance that they will be silenced by methylation or lost via recombination.
As the site of integraton is known, only a single line of "transgenic" mice is required.

CD Genomics has already updated its technology platforms to mainstream NGS, long-read sequencing, and microarray instruments. We continue to strive to offer the same reliable services to pharmaceutical and biotechnology companies, as well as academia and government agencies, for all your sequencing or array needs. If you are interested, please contact us directly for assistance.