Biomedical subjects

Keith Crouch

Publications and source records attributed to Keith Crouch.

4 recordsLinked to original sources

PubMed2006

An annotated cDNA library of juvenile Euprymna scolopes with and without colonization by the symbiont Vibrio fischeri.

BACKGROUND: Biologists are becoming increasingly aware that the interaction of animals, including humans, with their coevolved bacterial partners is essential for health. This growing awareness has been a driving force for the development of models for the study of beneficial animal-bacterial interactions. In the squid-vibrio model, symbiotic Vibrio fischeri induce dramatic developmental changes in the light organ of host Euprymna scolopes over the first hours to days of their partnership. We report here the creation of a juvenile light-organ specific EST database. RESULTS: We generated eleven cDNA libraries from the light organ of E. scolopes at developmentally significant time points with and without colonization by V. fischeri. Single pass 3' sequencing efforts generated 42,564 expressed sequence tags (ESTs) of which 35,421 passed our quality criteria and were then clustered via the UIcluster program into 13,962 nonredundant sequences. The cDNA clones representing these nonredundant sequences were sequenced from the 5' end of the vector and 58% of these resulting sequences overlapped significantly with the associated 3' sequence to generate 8,067 contigs with an average sequence length of 1,065 bp. All sequences were annotated with BLASTX (E-value < -03) and Gene Ontology (GO). CONCLUSION: Both the number of ESTs generated from each library and GO categorizations are reflective of the activity state of the light organ during these early stages of symbiosis. Future analyses of the sequences identified in these libraries promise to provide valuable information not only about pathways involved in colonization and early development of the squid light organ, but also about pathways conserved in response to bacterial colonization across the animal kingdom.

Aliivibrio fischeri↗

PubMed2004

High-throughput gene discovery in the rat.

The rat is an important animal model for human diseases and is widely used in physiology. In this article we present a new strategy for gene discovery based on the production of ESTs from serially subtracted and normalized cDNA libraries, and we describe its application for the development of a comprehensive nonredundant collection of rat ESTs. Our new strategy appears to yield substantially more EST clusters per ESTs sequenced than do previous approaches that did not use serial subtraction. However, multiple rounds of library subtraction resulted in high frequencies of otherwise rare internally primed cDNAs, defining the limits of this powerful approach. To date, we have generated >200,000 3' ESTs from >100 cDNA libraries representing a wide range of tissues and developmental stages of the laboratory rat. Most importantly, we have contributed to approximately 50,000 rat UniGene clusters. We have identified, arrayed, and derived 5' ESTs from >30,000 unique rat cDNA clones. Complete information, including radiation hybrid mapping data, is also maintained locally at http://genome.uiowa.edu/clcg.html. All of the sequences described in this article have been submitted to the dbEST division of the NCBI.

Animals↗

PubMed2004

1274 full-open reading frames of transcripts expressed in the developing mouse nervous system.

As part of the trans-National Institutes of Health (NIH) Mouse Brain Molecular Anatomy Project (BMAP), and in close coordination with the NIH Mammalian Gene Collection Program (MGC), we initiated a large-scale project to clone, identify, and sequence the complete open reading frame (ORF) of transcripts expressed in the developing mouse nervous system. Here we report the analysis of the ORF sequence of 1274 cDNAs, obtained from 47 full-length-enriched cDNA libraries, constructed by using a novel approach, herein described. cDNA libraries were derived from size-fractionated cytoplasmic mRNA isolated from brain and eye tissues obtained at several embryonic stages and postnatal days. Altogether, including the full-ORF MGC sequences derived from these libraries by the MGC sequencing team, NIH_BMAP full-ORF sequences correspond to approximately 20% of all transcripts currently represented in mouse MGC. We show that NIH_BMAP clones comprise 68% of mouse MGC cDNAs > or =5 kb, and 54% of those > or =4 kb, as of March 15, 2004. Importantly, we identified transcripts, among the 1274 full-ORF sequences, that are exclusively or predominantly expressed in brain and eye tissues, many of which encode yet uncharacterized proteins.

Animals↗

PubMed2003

EST-based gene discovery in pig: virtual expression patterns and comparative mapping to human.

A molecular understanding of porcine reproduction is of biological interest and economic importance. Our Midwest Consortium has produced cDNA libraries containing the majority of genes expressed in major female reproductive tissues, and we have deposited into public databases 21,499 expressed sequence tag (EST) gene sequences from the 3' end of clones from these libraries. These sequences represent 10,574 different genes, based on sequence comparison among these data, and comparison with existing porcine ESTs and genes indicate as many as 4652 of these EST clusters are novel. In silico analysis identified sequences that are expressed in specific pig tissues or organs and confirmed the broad expression in pig for many genes ubiquitously expressed in human tissues. Furthermore, we have developed computer software to identify sequence similarity of these pig genes with their human counterparts, and to extract the mapping information of these human homologues from genome databases. We demonstrate the utility of this software for comparative mapping by localizing 61 genes on the porcine physical map for Chromosomes (Chrs) 5, 10, and 14.

Algorithms↗