Research Programs

Costs & Basic Guidelines for Other Services

Tail Snips of Potential Transgenic Pups
Cryopreservation of Mouse Embryos
Electroporation of Embryonic Stem Cells

Tail Snips of Potential Transgenic Pups

The facility provides tail snips of potential transgenic pups at 7-10 days of age as part of the micro-injection project. We are now prepared to isolate DNA from those tail snips, and provide 500 ul of clean DNA suitable for PCR or Southern analysis. Although we do not routinely quantify DNA isolated by our procedures, 2-10 ul has proven suitable for PCR analysis. The PCR analysis done by the facility is performed one time, with the lab providing primer sequences and parameters for analysis previously worked out by the lab. A typical injection project would involve analysis of 20-30 pups.

Procedure	Costs	Goals
Tail DNA Extraction/Purification	$35/up to10 tails $35/each additional 10 tails	500 ul of purified DNA
Above, plus PCR Analysis	$125/1-10 samples $125/each additional 10 samples	identification of positive samples

Cryopreservation of Mouse Embryos:

Either slow freezing using a ramp freezer or quick freezing by vitrification of 8 cell to morula stage embryos in order to preserve an unused line for future use, or to protect against loss of a line.

The laboratory will provide at least 5 adult male mice, 2-8 months of age, either proven fertile or experienced breeders. The facility will purchase 10-15 young females of the same or a closely related strain, to superovulated and mate with the males. 2.5 day p.c. embryos will be harvested and frozen. The total number of embryos frozen will be determined empirically by doing an initial test freeze, immediately followed by a test thaw. The percentage of embryos that survive thaw, and then go on to implant and develop into live pups will establish the parameters of the cryopreservation project. The laboratory will be responsible for testing the genotype of the pups resulting from the test thaw.

Total costs may vary if the line proves difficult to freeze, if the investigator desires more than the normal number of embryos frozen, or if the background strain chosen is more expensive to purchase or difficult to superovulate. If the line proves resistant to cryopreservation, ½ the cost will be assessed to cover the cost of the attempt.

Procedure	Cost	Parameters	Experimental Goals
Freeze	$500	150-200 embryos frozen, one test thaw performed	Preservation of line
Thaw	$200	20 embryos thawed, transfer surgery, tail sampling	5-8 live mice
Storage	$50	Annual storage fee in Transgenic Facility: SPACE IS LIMITED AT THIS TIME Storage is in cryovials or straws (special straw holders that fit in canes will be provided by the facility).	--

Electroporation of Embryonic Stem Cells:

Embryonic stem cells currently must be purchased or obtained in a collaborative agreement by the laboratory. Three commercial vendors are linked off this site under ES cell vendors. The facility requests as much information as possible on the growing conditions for the cells be provided. The electroporation request form has the details for DNA preparation and amount required. The facility will electroporate the ES cells with the DNA provided; will plate the cells, add selection (positive or positive/negative; we use G418 for neomycin selection, and gancyclovir for negative selection), and pick 150-300 clones. These clones will be replica-plated in 96 well plates; two copies will be frozen, and one will be grown off feeders to provide cells for DNA analysis. Expansion to 24 well plates is provided at an extra charge. The laboratory will identify positive clones, within 1 month. Plates are stored at -80, and delay in identifying clones may result in loss of viability of ES cell clones. The facility will expand up to 5 positive clones; freezing back 3 vials of cells per clone in liquid N2. More clones can be expanded and frozen at an extra charge. The frozen vials will be turned over to the laboratory for storage, unless they are scheduled for microinjection.

Procedure	Cost	Parameters
Electroporation	$4000	150-300 clones picked, up to 5 positive clones expanded
Expansion to 24 well plates	$500	Clones frozen in 96 well plates; 24 well samples for DNA analysis
Expansion of positive clones (more than 5)	$20/clone	Clone expanded from 96 well plate; 2-3 vials frozen per clone

This page was last updated on: January 19, 2007.