Meeting Abstract

71.2  Saturday, Jan. 5  NGS analyses of genes expressed during limb regeneration in the crab, Uca pugilator DAS, S.*; NAJAR, F.Z.; LAI, H.C.; WILEY, G.; GAFFNEY, P.M.; ROE, B.A.; DURICA, D.S.; Univ. of Okahoma; Univ. of Okahoma; Univ. of Okahoma; OMRF; OMRF; Univ. of Okahoma; Univ. of Okahoma sunetra.das-1@ou.edu

Limb regeneration in fiddler crabs involves formation of a proliferating blastema, its differentiation into a segmented ‘mini limb’ (basal phase) and hypertrophic growth of the ‘mini limb’ via protein synthesis and water uptake (proecdysial phase). These phases are tightly coordinated with the molt cycle, i.e. accompanied by distinct fluctuations in circulating ecdysteroid titers, providing a useful model system to study changes in global gene expression. Among crustaceans, genomic and RNA-seq datasets are limited. We gave generated pilot transcriptome profiles to examine steady state changes in global gene expression during the limb regeneration process, including blastema, early and late proecdysial limb buds, using 454 and Illumina sequencing technology (NGS). Following sequencing, the reads were assembled de-novo by using the Newbler Assembler for 454 and the Trinity and SOAP Assemblers for Illumina sequence data. We have generated 103,700 and 704,395 sequences (all libraries combined) from the 454 and Illumina platforms, respectively. The average contig lengths from proecdysial libraries built by the three assemblers were: 511 bp (Newbler), 186 bp (Trinity) and 629 bp (SOAP). Analyses of the sequence data are available online at http://www.genome.ou.edu/crab.html, where the databases are both BLAST and keyword searchable. The database contains putative isoforms not detected through cDNA library cloning or anchored PCR. We have also obtained metabolic profiles from early blastemal, and early and late proecdysial limb buds using the KEGG database. Further analysis of metabolic profiles, in association with experimental manipulation of ecdysteroid responsiveness, should provide information on gene pathways subject to ecdysteroid control.