Abstract
BackgroundBiologists 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.ResultsWe 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).ConclusionBoth 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.
Highlights
Biologists are becoming increasingly aware that the interaction of animals, including humans, with their coevolved bacterial partners is essential for health
A total of 11 cDNA libraries (Table 1) were generated from 5 pools of squid light organ RNA, containing clones with insert sizes ranging from 0.35 to 2.5 kb and an average length of ~1 kb. From these cDNA libraries, sequencing generated a total of 42,564 3' expressed sequence tags (ESTs) sequences with an average sequence length of 692 bp and an average phred value of 39.7
A histogram illustrating the total number of unique E. scolopes ESTs, the total number of ESTs annotated as a known protein with an E-value threshold of e-03, the total number of ESTs annotated with at least one category of Gene Ontology (GO) and the number of genes annotated in each of the 3 major GO categories, biological process, molecular function and cellular component
Summary
Biologists are becoming increasingly aware that the interaction of animals, including humans, with their coevolved bacterial partners is essential for health. Large-scale sequencing efforts are allowing biologists to define the communities associated with particular animal species (e.g., [2,3,4,5,6]), and development of such methods as microarray analyses are enabling biologists to determine host and symbiont responses under natural and experimental conditions [6,7] In addition to these technological advances, the development of powerful models for the study of animal-bacterial interactions has been essential. The study of a wide variety of models will shed light on evolutionarily conserved mechanisms as well as the basis of diversity To this end, two principal, complementary model types are under investigation: those using germ-free or gnotobiotic animals and binary associations. I.e., those composed on one host species and a population of one microbial species, occur most often in invertebrate animals and offer relatively simple experimental systems for the study of animal-microbe relationships [11,12,13]
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