TOWLE, D.W.*; SMITH, C.M.; Mt. Desert Island Biological Laboratory: Expressed sequence tags in a normalized cDNA library prepared from multiple tissues of the American lobster Homarus americanus

Although the American lobster Homarus americanus provides a highly valuable fishery in northern New England and the Atlantic provinces of Canada, populations in other areas are under stress. In an effort to identify gene expression patterns that may allow an assessment of stress levels in animals sampled from the field, total RNA was isolated separately from ten different tissues of adult intermolt lobsters: gill, epipodite, branchiostegite, heart, ovary, testis, antennal gland, abdominal muscle, hepatopancreas, and brain. Following quality analysis, equal amounts of total RNA were pooled for construction of a mixed tissue cDNA library. Normalization reduced the abundance of arginine kinase encoding cDNAs from 2.47% in the original library to 0.0465% in the finished product, a 53-fold reduction. Clone isolation and 5�-end sequencing of 5,568 plasmid inserts produced 4,604 dbEST-submissable sequences following processing by trace2dbest software. Average length of the expressed sequence tags was 572 nucleotides and the average length of high quality sequence was 453. Clustering by partigene software revealed 1,412 ESTs in 579 clusters plus 3,194 singletons for a total yield of 3,773 unique sequences. Approximately 50% of these sequences returned a high-scoring hit with blastx. Results are available in dbEST at NCBI and at www.marinegenomics.org. Supported by the Maine Biomedical Research Infrastructure Network (NCRR 1P20RR16463-01).