An Exploratory Study of Functional Genes Related to Nutrient Uptake in an Upwelling-Representative Diatom Using Transcriptome Sequencing---Gene Expressions in Purified Cultures and Field Measurements in the East China Sea

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Project title

Code/計畫編號

NSC99-2313-B019-002

Translated Name/計畫中文名

以全轉錄體定序探索湧升區代表性矽藻中之營養鹽吸收功能性基因---純化藻種之基因表現與東海實測

Project Coordinator/計畫主持人

Jeng Chang

Funding Organization/主管機關

National Science and Technology Council

Department/Unit

Institute of Marine Biology

Website

https://www.grb.gov.tw/search/planDetail?id=2103317

Year

2010

Start date/計畫起

01-08-2010

Expected Completion/計畫迄

01-07-2011

Bugetid/研究經費

1000千元

ResearchField/研究領域

生物技術(農)
漁業

Description

Abstract

"矽藻是東海湧升流海域重要的初級生產者，而監測指標基因表現量是瞭解矽藻如何因應營養鹽濃度與比例變化的有效方法。目前選用的矽藻營養鹽指標有偵測氮與磷限制的硝酸鹽傳輸蛋白和磷酸鹽傳輸蛋白，但面臨指標基因種類偏少以及在海洋中不易選殖同源基因的等困境。有鑑於此，本申請案計劃針對一種在東海湧升流海域常見的矽藻 -- 相關角刺藻 -- 進行全轉錄體定序，也就是大規模的決定藻細胞中所有 mRNA 的序列。經由比較各種營養鹽濃度及比例下產生之全轉錄體，就能夠快速且大量的搜尋營養鹽吸收相關基因，並自其中挑選表現量可忠實反應營養鹽缺乏程度者做為指標基因。具有潛力的基因將先用實驗室純種培養的藻種進行確認，同時也要在東海中調查同源基因的序列岐異度。由於角刺藻是湧升流海域的優勢種類，以相關角刺藻序列進行海洋基因選殖可提高成功率。另外也將建立相關角刺藻的基因組基因庫，將搜尋自全轉錄體的基因片段擴展為基因全長序列。根據這些過程將設計出一套適用的指標基因，並使用定量聚合脢鎖鏈反應偵測湧升流海域矽藻的生理狀態。本申請案採用了先進的技術研究具生態意義的議題，預期所產生的結果將能對漁業資源、海洋優養化、以及二氧化碳的吸收提供重要資訊。" "Marine diatoms are important primary producers in a shelf break upwelling system of the East China Sea. To understand how diatoms adapt to the ever changing environment in terms of nutrient concentrations and ratios, a good method is to monitor the expression of indicator genes. The diatom indicator genes currently in use include the nitrate transporter gene for nitrogen deficiency and the phosphate transporter gene for phosphorus deficiency. However, the numbers of suitable indicator genes are still few, and genes homologous to laboratory-defined indicators are difficult to obtain in marine environments. For these reasons, this application proposes to generate a transcriptome data bank for a common diatom in the upwelling region, Chaetoceros affinis, which involves a large-scale sequencing of all mRNA produced in algal cells. Via the comparisons between transcriptomes generated under various nutrient concentrations and ratios, genes related to the utilization of ammonium, nitrate, and phosphate can be rapidly and thoroughly searched, and genes with an expression that faithfully reflects the degree of nutrient deficiency will be selected as an indicator gene. The performances of potential indicator genes will be confirmed using laboratory cultures. At the same time, the diversity of their homologous sequences in the East China Sea will be investigated. Since species in Chaetoceros genus are one of the dominant groups in the upwelling region, gene amplification in this region using C. affinis sequences should enjoy higher successful rates. In addition, a genomic DNA library will be constructed for C. affinis, which can be used to obtain full-length sequences of interested gene fragments found in transcriptome analysis. These results will allow the design of a series of indicator genes which can be applied to natural environments, and the method of quantitative polymerase reaction will be used to detect the expression level of these genes in the upwelling region. This proposal employed advanced techniques to study an ecologically important topic. Technology and experimental results generated by this project will provide important information to the management of fishery resources, the control of marine eutrophication, and the absorption and sinking of carbon dioxide."

Keyword(s)

東海
浮游植物
營養鹽缺乏
指標基因
全轉錄體
矽藻
East China Sea
phytoplankton
nutrient stress
marker genes
transcriptome
diatom