Suppression subtractive hybridization PCR isolation of cDNAs from a Caribbean soft coral
Jose V. Lopez*1,2,3 · Angela Ledger2 · Lory Z. · Santiago-Vázquez3,4 · Mihai Pop5 · Dan D. Sommer5 · Llanie K. Ranzer3,6 · Robert A. Feldman7 · Russell G. Kerr3,8
Center, Nova Southeastern University, Dania Beach, FL, USA
*Corresponding author: firstname.lastname@example.org
Financial support: The authors gratefully acknowledge financial support from Florida Sea Grant College (R/LR-MB-23 to JVL and R/LR-MB-14 to RK). This material is also based upon worked supported by the National Science Foundation under a grant awarded to L.Z. Santiago-Vázquez (awards #0310283 and 0514500). MP and DDS were supported in part the NIH grant R01-HG-0004885 to MP.
Keywords: cDNA, Erythropodium caribaeorum, EST, Gorgonia ventalina, gorgonian, sea fan.
Transcriptomic studies of marine organisms are still in their infancy. A partial, subtracted expressed sequence tag (EST) library of the Caribbean octocoral Erythropodium caribaeorum and the sea fan Gorgonia ventalina has been analyzed in order to find novel genes or differences in gene expression related to potential secondary metabolite production or symbioses. This approach entails enrichment for potential non-“housekeeping” genes using the suppression subtractive hybridization (SSH) polymerase chain reaction (PCR) method. More than 500 expressed sequence tags (ESTs) were generated after cloning SSH products, which yielded at least 53 orthologous groups of proteins (COGs) and Pfam clusters, including transcription factors (Drosophila Big Brother), catalases, reverse transcriptases, ferritins and various “hypothetical” protein sequences. A total of 591 EST sequences were deposited into GenBank [dbEST: FL512138 - FL512331, and GH611838]. The results represent proof of concept for enrichment of unique transcripts over housekeeping genes, such as actin or ribosomal genes, which comprised approximately 17% of the total dataset. Due to the gene and sequence diversity of some ESTs, such sequences can find utility as molecular markers in current and future studies of this species and other soft coral biogeography, chemical ecology, phylogenetics, and evolution.