Pronuclear Injection

Hybrid and FVB inbred strains

Microinjection of purified DNA constructs for the production of transgenic mice. This includes injection of a single transgene construct (prepared by the investigator). A minimum of two transgenics or twenty weaned animals (or embryos), whichever comes first, will be prepared for each set of injections charged to an investigator.

Additional days of injection for hybrids and FVB inbreds: The standard fee covers up to 2 microinjection days. Any additional days of injection that are required will incur a daily fee.

C57Bl/6 and other common inbred strains

The use of inbred strains other than FVB will be assessed a surcharge fee with a maximum of 3 days of microinjection.

Additional days of injection for C57Bl/6 and other inbreds: The use of common inbred strains that require the surcharge fee will cover 3 days of injection, and subsequent injection days will be subject to an extra daily fee.

Microinjection into less commonly available, more expensive strains

Special requests for microinjection into more costly strains of mice will be subject to a strain surcharge plus the purchase cost of the animals.

Additional days of injection (beyond 3) into strains that are not commonly available will be assessed a daily fee plus the purchase cost of the animals.

ES cell Injection

Microinjection of selected embryonic stem (ES) cell clones to establish chimeric mice for the production of "knockout" mouse lines. The fee covers the injection of a single selected ES cell clone and reimplantation of a minimum of 20 blastocysts (two surgeries).

The standard fee covers injection of 129/Sv derived ES cells into C57Bl/6 blastocysts. Injection of ES cells derived from other strains (such as C57 Bl/6 ES cells) is much less efficient, and requires the purchase of different host strains. Requests to produce chimeric mice from injections of ES cells derived from mouse strains other than 129/Sv will be subject to an additional surcharge.

Rederivation

Rederivation of mouse lines infected with various mouse pathogens to allow these lines to be shipped to collaborators at other institutions or to allow them to be moved into cleaner, barrier facilities at Mount Sinai. The fee covers the cost to embryo rederive each line, using C57Bl/6, FVB/N, or F1 hybrid strains.

Other mouse strains (such as 129/Sv or Balb/c) are very difficult to use for embryo rederivation. The use of strains other than C57Bl/6, FVB/N, or F1 hybrids will result in an additional surcharge for the rederivation.

Some investigators may request rederivations using females provided from their research colony. This is typically done in the case where breeding with wild-type females is undesirable for maintaining certain genetic markers. An example of this is a line of double knockouts. Embryo rederivations using investigator provided transgenic or knockout females may not work, and the MGSRF will still assess the standard fee even if the rederivation attempts fail.

Sperm Cryopreservation/Recovery

The Mouse Genetics SRF has established a cryopreservation program for the long-term storage of mouse sperm to reduce the need for constant maintenance of lines which are not essential for current research needs.

Two levels of service for sperm cryopreservation will be offered:

Basic Sperm Cryopreservation

Sperm will be isolated and cryopreserved from a single, genotyped male. Other than visual observation of sperm density and motility, no further assessments will be made of the viability of the cryopreserved sperm. This level of service should be used for mouse lines which are not part of an active research program, and which will most likely not need to be recovered at some point in the future.

Sperm Cryopreservation Plus

Sperm will be isolated and cryopreserved from a single, genotyped male. In addition to visual observation of sperm density and motility, in vitro fertilization (IVF) of wild-type eggs will be performed to assess the viability of the frozen sperm. It is important to note that the eggs used for this IVF are derived from F1 hybrid lines. Attempts to recover lines in the future using eggs from inbred strains may yield very different IVF efficiencies. This level of service should be used for mouse lines which are part of an active research program, or which are likely to be recovered at some point in the future. Investigators should be certain that the sperm frozen from an important line is capable of fertilizing eggs before they substantially reduce or eliminate the line.

IVF Recovery

Upon request, Mouse Genetics SRF will recover lines from which sperm has been frozen. This requires IVF followed by recovery surgeries of fertile eggs into pseudopregnant female mice.

Standard IVF

Sperm from hybrid or FVB inbred mice will be used in standard IVF assays to generate 2-cell embryos for the recovery of cryopreserved lines

Zona-drilling IVF

Sperm from inbred lines, such as C57Bl/6, which do not perform well in standard IVF assays, will be used for IVF with eggs that have been micromanipulated to introduce small holes in the zona pellucida. This procedure significantly improves the yield of 2-cell embryos for recovery surgery.