Wild Cordyceps Sinensis CDNA Library Construction, EST Sequencing And Functional Genes Analysis

The Cordyceps sinensis is one of the most famous Chinese traditional medical herbs. Expressed sequence tags (ESTs) can obtain detailed information concerned on molecular level and supply the Cordyceps sinensis'genomic structure and the information of many genes. Therefore,it is a significant case study valuable for the furtuer achievement .The collections of Cordyceps sinensis'ESTs data have great contribution in discovery of new genes and preparation of gene chip . In this study,cDNA libraries were constructed from the stroma and sclerotium , respectively. Large-scale cDNA sequencing was performed and we had a first insight into the transcriptome of Cordyceps sinensis.Randomly picked,partially sequenced from their 5' end,21 845 ESTs were sequenced. All sequences were screened for quality using base-quality scores,low-quality ends of the sequence traces were removed.Short sequences that length less 100 bp and contaminant sequences were also removed.At last,we obtained 20 193 high-quality ESTs with an average sequence length of 545 bp for further analysis , which were assembled into 6481 consensus sequences(unisequences). The largest unisequence was composed of 545 ESTs,while the longest one was 1669 bp long.We obtained lots of information about its functional genes. By the six frame translations of the EST sequences , BLASTX analysis identified 4061 unisequences,which had significant similarities (E-value﹤10-5) to sequences deposited in the NCBI ' non-redundant protein database(NR,download from NCBI Jan 4th ,2006). However,less half of the unisequences showed similarities to known protein sequences or motifs, almost all of which were not previously described in this kind of fungi. A search against the NCBI non-redundant nucleotide sequences database(NT,download from NCBI Jan 4th ,2006) using the BLASTN algorithm revealed 1945 with high or moderate similarities to plant pathogen fungal sequences. Approximately 52.77% of the identities were to hypothetical proteins and proteins with unknown function.The other unisequences presented no matches with existing sequences in public databases. As a result of Gene Ontology (GO) project annotation,874 unisequences (3540 ESTs,17.53%) were classified into biological processes of the GO structure,involved in intracellular transport,primary metabolism,along with transcription and RNA processing,DNA replication,protein biosynthesis,proteolysis,response to oxidative stress,as prevalent processes.We analyzed these genomic data and characterized for potential functions, tissue-specific patterns of expression,and Cluster analysis of ESTs revealed major differences between the expressed gene content of stroma and sclerotium. The sequence data described are not available in a searchable public database.We detected glycosyl hydrolases,proteases,as well as serine and cysteine protease,which were predicted to encode secreted proteins that could function in virulence. Some of these cDNAs were more similar to fungal proteins than to other eukaryotic proteins. Conclusion: We provide a first glimpse into the gene content of Cordyceps sinensis. These resources will greatly promote research on this fungus.