We propose to create a needed resource for the molecular biology community, viz. a collection of cDNA libraries spanning murine embryogenesis. Given the tiny amounts of material involved, we will use the polymerase chain reaction (PCR) to construct these libraries, as we have done for a variety of human embryonic tissues. For this purpose, we have established conditions to obtain PCR-based cDNA libraries with both the appropriate complexity and insert length using long-PCR protocols that were originally applied to genomic DNAs. Using these techniques, we will construct cDNA libraries at each half-day of embryogenesis, thereby yielding 40 complex cDNA libraries (each with at least 10/6 independent primary clones). Additional complex cDNA libraries from unfertilized spermatocytes and oocytes, and from brain and heart from day 9 of development and daily thereafter until day 20 (time of birth) will also be constructed. We describe PCR-based methods to screen these libraries efficiently for transcription and to isolate longer transcripts. The availability of these libraries from Research Genetics will provide the material necessary to other researchers for a host of studies relating to development.
The cDNA libraries spanning murine development we will create constitute a desired and needed resource of interest to many laboratories. These libraries will be constructed and screened using novel PCR-based technologies. Once constructed, they will be a useful resource to a large number of investigators who will purchase the libraries from Research Genetics.
