以表現標誌序列重組南美白蝦之轉錄基因體並比較不同組織間的基因表現

Abstract

南美白蝦 ( Litopenaeus vannamei ) 在蝦類養殖的地位日趨重要，進行其基因轉錄體的研究可了解該物種的基因組成，有助於對蝦生理、遺傳、育種、與病理研究等基礎領域，以因應對蝦繁養殖上所面臨的問題。EST (Expressed Sequence Tag) 是cDNA的部分定序序列，可提供轉錄基因的序列資訊及相對表現數量，對於尚未完成完整基因體的物種，藉由EST定序能得到大量蛋白質基因的資訊。本研究蒐集公開資料庫上的白蝦ESTs序列，共161,241筆資料，利用與白蝦在分類上相近且完成完整基因體定序解析資訊的水蚤 (Daphnia pulex) 及果蠅 (Drosophila melanogaster) 為參考，利用序列的同源關係，設計序列分析流程，組裝得到16,886筆contigs及20,515筆singletons，共37,401筆重組序列；經過資料庫比對的序列註解程序後，超過40%的重組序列可在non-redundant protein database及Pfam database找到相似的序列或模組特徵。 在原始EST序列資料中，92%的是來自眼柄、鰓、血細胞、肝胰臟、淋巴器官及神經索，此六個組織的EST數量皆超過20,000筆。上述重組序列，以其 EST來源組織的EST數為基因表現量，代表六個組織的基因表現概況。分析這六個組織以重組序列的Gene Ontology 註解資訊，發現眼柄、肝胰臟及淋巴器官的表現基因功能分類有較明顯集中（enrichment）的傾向，挑選其中差異較大的眼柄及肝胰臟，進一步分析兩組織的重組序列組成在KEGG pathway 的配置方式，是否有集中程度的差異。綜合 KEGG pathway 與 GO term 的分析結果顯示，兩組織間都有共同表現的重組序列，大多是參與轉譯過程的基因，眼柄組織特有序列較集中表現actin及myosin兩個基因，肝胰臟組織的特有序列，則是表現血藍素 (hemocyanin)。本研究提供不同的序列組裝策略，運用重組序列之註解了解組織特異性，其結果能幫助我們對白蝦的基因更加了解，過程得到的資料也可提供後續研究者作為研究之參考。 參考網站：http://ips.sinica.edu.tw/lv

The Pacific white shrimp, Litopenaeus vannamei, has became a more important animal in shrimp aquaculture. Although its genome has not been fully sequenced, this economically important species has attracted researchers for building a rich source of ESTs for benefiting both the basic biology and the applied science. Expressed Sequence Tags (ESTs), are short sequences derived from a batch-wise partial sequencing result of cDNA library, which may represents the relative expression level of transcripts in the library. It is an effective approach to get numerous sequences of protein coding genes of a species without previous knowing about the genome context. In this study, we collected 161,241 ESTs of white shrimp from public database. Reference sequences from two taxonomically closed species Daphnia pulex and Drosophila melanogaster were used for selecting subset for de novo assembling. Totally, 37,401 assembled sequences, including 16,886 contigs and 20,515 singletons, were obtained. Over 40% of the assemblies could be matched to homolog sequences in non-redundant protein database and/or protein domain feature in Pfam database. Six tissues including eyestalk, gills, hemocyte, hepatopancreas, lymphoid organ and nerve cord are the major tissue source of this collection. Assemblies were annotated and the expression level of each assembled contig/ singleton in each tissue was calculated by the number of ESTs contributed to this contig/singleton. Enrichment analyses were further applied to detect Gene Ontology terms, and KEGG pathway for describing the unique function of each tissue. For example, the unique expressing subset of hepatopancreas is related to hemocyanin in KEGG pathway enrichment analysis result, while the unique subset in eyestalk represent actin and myosin genes. In summary, this study provides an alternative strategy of assembling EST and the data produced from this research can assist any studies about white shrimp in the future. Reference URL: http://ips.iis.sinica.edu.tw/lv