Microinjections and Expected Results

Microinjections

Transgenic DNA is initially microinjected into approximately 150 to 200 fertilized single-cell embryos, which is generally enough embryos to yield 20 to 25 live births or five females at the desired day of gestation for transient transgenesis projects. Injected embryos are then surgically transferred into the oviduct of pseudopregnant females. Females are weighed at 10 days post surgery to assess pregnancy and relative size of litter. If problems are perceived at this time, additional injections are immediately scheduled.

Pups are typically born 19 days post surgical transfer. Occasionally, litters are lost due to cannibalism, which can be caused by any number of factors, including noise or irregular vibrations (i.e., from major construction projects). Pregnant females are assessed for their mothering instincts, such as nesting, just prior to the expected delivery date and foster mothers (nursing moms) are often available if problems arise. If litters are lost, additional injections are scheduled immediately.

Expected Results

• Transgenesis rates of between 10 to 25 percent, or two to seven founders, are expected for most constructs
• If no founders are identified, a second set of injections will be performed
• Consultation and Southern blot analysis (if PCR is used for initial transgenesis screen) is required, however, prior to a second round of injections