Geisha: A Gallus (chicken) EST and in situ hybridization approach for identifying genes expressed during early embryogenesis
AuthorBell, George W.
AdvisorAntin, Parker B.
MetadataShow full item record
PublisherThe University of Arizona.
RightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
AbstractWith ever-increased quality and quantity of vertebrate genomic sequences, gene finding remains problematic, and predicting gene function from sequence remains a tremendous challenge. Nevertheless, similarity of sequence and patterns of gene expression are rapid methods for providing insight into potential functional roles of novel genes. For developmental studies, microarrays offer a means for screening a large number of sequences for expression within defined tissues and/or organs, but each experiment profiles expression level for only one anatomical region at one specific developmental stage. Whole mount in situ hybridization (ISH) offers an alternative approach that gives precise spatial and temporal resolution of gene expression throughout an embryo. I predicted that combined high throughput analysis of expressed sequence tags (ESTs) and whole mount ISH analysis of novel clones would effectively identify novel developmentally regulated avian genes. 5' ESTs were generated from randomly chosen cDNA clones of a chick late gastrula endoderm-mesoderm library. Following screening to remove ubiquitously expressed clones, internal clustering, and comparison to GenBank sequences, remaining cDNAs (both similar and dissimilar to known genes) were screened for expression in HH stage 4-12 embryos by automated high throughput whole mount ISH. Comparison to GenBank sequences by blastn and blastx (e < 0.05) revealed that one quarter of all 955 ESTs represented novel genes. Of these novel clones, ISH showed that about one quarter (60 clones) exhibited patterned expression during embryogenesis. EST sequences, ISH images and corresponding blast reports were assembled into a freely accessible Web database at http://geisha.biosci.arizona.edu.
Degree ProgramGraduate College