The expression of the CRF1 gene in mouse embryonic development

A mouse gene cloned in the laboratory of James and Olivia Smith at Baylor College of Medicine was found to be involved in motor function. This gene was named CRFl for Clq-Related Factor 1, and using in situ hybridization, it was found to be expressed mainly in the brainstem of adult mice. There is a gene similar to the CRFl gene in humans. By determining this gene's role in mouse development, more will be known about the development of human motor function. To determine the stage of embryonic development at which the expression of this gene begins, a series of in situ hybridization experiments were performed on saggital sections of embryos at 11.5, 12.5, 13.5, 14.5, and 16.5 days of development using a diglabeled riboprobe made from CRF template DNA. Two types of substrate detection, NBT/BCIP and fluorescence, were used to visualize the probe. In several experiments, no specific hybridization occurred. In the remainder of the experiments, the hybridization that occurred in specific tissues was not repeatable and therefore unreliable. Introduction In recent years, we have been able to discover many new genes and their functions. Many times genes are discovered in animals that are related to genes in humans. We are very interested in finding out what these new genes do in humans and what abnormalities and diseases they may cause. The first step in discovering these genes' roles in human life is finding out what the related gene does in animals' lives. Many genes have been found that play a role in development. These genes determine an organism's body type, musculature, physical characteristics, and so forth. If these genes have a role in human development, we will begin to understand how we are formed into such complex beings. The gene that the Smith lab discovered is one such gene. The CRF 1 gene was found in mice, and humans have a gene that is related to it. This gene was determined to be important in motor function of adult mice. Without it, mice would not be physically capable animals. The implications on human life are enormous. By discovering when and where this gene is expressed in humans, we may be able to learn more about our own motor abilities and how our muscles and nerves work together to enable us to perform complex motor tasks. By first determining when and where this gene is expressed in mice, we can begin to answer some of these questions about humans. A laboratory technique called in situ hybridization allows us to determine this. In situ hybridization, from Latin for "in site", allows us to determine where in the body a particular gene is expressed. A probe is made of genetic material that is identical or similar to the gene in question. It is usually labeled with an enzyme or radioactivity that is easily observed. By then applying the probe to a tissue, it is possible to see if the gene is expressed at that site. If a series of tissues are used that are at different stages of development, it would also be possible to determine when the gene is expressed--whether during embryonic development or after birth. Materials and Methods Embryo Preparation The embryos must be prepared so they stay fresh and can be used in many experiments. The tissue is embedded in paraffin wax so that it remains intact when