2008 Annual Report
1a.Objectives (from AD-416)
Sequencing and profiling of functional transcripts constructed in expressed sequence tags (ESTs) of Coptotermes formosanus to fulfill the following objectives: a) Identifying and annotating of genes specifically associated with post-embryonic polyphenism (caste differentiation and development); b) Discovering of genes specifically responding to environmental cues and internal signals (pesticides, food sources, juvenile hormones, etc.); and c) Characterizing of genes uniquely involved in critical physiological pathways (digestion, molting, immunity, etc.). The fulfillment of these objectives would directly lead to achieving the following goals: a) providing biochemical/physiological/molecular bases for disruption of colony formation, development and survival; b) Discovering novel target site(s) that would be developed into new control strategies that could be incorporated into effective area-wide integrated management of Formosan subterranean termites.
1b.Approach (from AD-416)
A cDNA library representing expressed genes in each different developmental stage of Coptotermes formosanus will be constructed. To facilitate transcriptome analysis and rare gene discovery, repeated transcripts will be proportionally removed from the cDNA library using the procedure of cDNA normalization. The cDNA library would contain approximately 400,000 independent clones, an estimate of at least 16X coverage of the entire expressed genes, provided that the protein-coding capacity of invertebrate genomes is in the range of 16,000 to 25,000 genes. The sequencing and gene data assembly of the cDNA library will be cooperatively conducted in JCVI. Eighty thousand clones will be initially sequenced at both 5’- and 3’-ends. The resulting EST sequences will be compared against existing databases and annotated using the Basic Local Alignment Search Tool (BLAST). Batches of sequences will be sequentially released to GenBank for public access. Unique genes and singletons would be selected for array and gene expression analysis. Differentially-expressed genes or development-stage specific genes will be preferentially analyzed quantitatively (such as real-time PCR) or qualitatively (such as gene silencing by RNA interference).
J. C. Venter Institute has completed cDNA sequencing on approximately 150,000 clones obtained using template Ribonucleic Acid (RNA) from virtually all life stages of the Formosan Subterranean termite, Coptotermes formosanus. Eighty-eight percent of the sequence reads were judged to be of good quality with an average read length of 555 base pairs. Sequence identities were found to include genes from C.formosanus, other termite species, other insect species and protozoan symbionts known from C. formosanus. Diribonucleic acid (DNA) sequence similarity was found to include genes from C. formosanus, other termite and insect species, and protozoan microorganisms known to be found in the gut of C. formosanus. Some genes of interest identified through sequence homology are lipopolysaccharide binding protein, trypsin binding protein, chitin binding protein, an actin binding protein, a zinc binding dehydrogenase among others. Further analysis is being completed to determine other sequence homologies and to determine possible gene function. Plans are underway for further sequencing to provide complete genome coverage of the termite and its gut symbiotic protozoans. We will then perform analysis to determine termite caste and colony growth and differentiation associated genes in order to determine possible biologically-based control measures for the termite. Progress of this project is monitored by phone calls, e-mails, teleconferences and regular mail.