環保專案成果報告查詢系統

專案計畫編號
經費年度	113
計畫經費	2,450 千元
預算科目	提升土壤及地下水污染調查及整治技術工作
專案開始日期	2024/03/06
專案結束日期	2024/12/31
專案主持人	賴政佑
主辦單位	國環院檢測技術中心
承辦人	蔡志賢
執行單位	益農有限公司
專案分類	土壤及地下水、環境監測及檢測
中文關鍵字	第三代微生物總基因體定序、重金屬總基因體關聯性分析、分子檢測
英文關鍵字	Third Generation Metagenome Sequencing, Metagenome-wide association study with heavy metal, Molecular diagnostics
協同主持人
共同主持人
計畫聯絡人	余中鼎
計畫聯絡電話	0920141249
計畫聯絡信箱	jongding.yu@inong.com.tw

中文摘要	本研究以創新的“先統計，後設計”分子檢測開發策略，透過第三代微生物總基因體分析技術，首先量化底泥樣品中的微生物重金屬抗性基因分布，透過基因數據與重金屬濃度數據，並考量環境氣候因子進行關聯性統計分析（ＭＷＡＳ），遴選出最佳重金屬代表性基因以開發反應重金屬濃度的引子探針，引子探針設計上考量基因功能序列特性以設計出更精準之重金屬ｑ－ＰＣＲ篩檢引子及探針，最後完成ｑ－ＰＣＲ篩檢引子及探針測試並評估其可行性。於彰化縣灌溉渠道番雅溝及新埤舊圳採集底泥樣品完成重金屬檢驗及微生物族群總基因體分析，並納入先前「微生物菌相、訊號基因與訊號因子檢測鑑定分析」案之底泥樣品重金屬檢測及微生物族群總基因體分析資料，完成微生物重金屬抗性基因量化分析。由基因量化分析數據與重金屬濃度數據的關聯性分析結果進一步遴選出3組具專一性之管制重金屬代表性基因：鉻ｃｚｃＣ、鉻ｚｎｅＢ／ｈｍｘＢ及鋅ｃａｄＡ／ｙｖｇＷ，進一步設計共38組引子及探針進行ｑ－ＰＣＲ測試，測試結果從鉻的設計引子中選出Ｃｒ＿ｚｎｅＢｈｍｘＢ＿５以及從鋅的設計引子中選出Ｚｎ－ｃａｄＡｙｖｇＷ＿３１作為適用性最高的引子對。
中文關鍵字	第三代微生物總基因體定序、重金屬總基因體關聯性分析、分子檢測

中文摘要

本研究以創新的“先統計，後設計”分子檢測開發策略，透過第三代微生物總基因體分析技術，首先量化底泥樣品中的微生物重金屬抗性基因分布，透過基因數據與重金屬濃度數據，並考量環境氣候因子進行關聯性統計分析（ＭＷＡＳ），遴選出最佳重金屬代表性基因以開發反應重金屬濃度的引子探針，引子探針設計上考量基因功能序列特性以設計出更精準之重金屬ｑ－ＰＣＲ篩檢引子及探針，最後完成ｑ－ＰＣＲ篩檢引子及探針測試並評估其可行性。於彰化縣灌溉渠道番雅溝及新埤舊圳採集底泥樣品完成重金屬檢驗及微生物族群總基因體分析，並納入先前「微生物菌相、訊號基因與訊號因子檢測鑑定分析」案之底泥樣品重金屬檢測及微生物族群總基因體分析資料，完成微生物重金屬抗性基因量化分析。由基因量化分析數據與重金屬濃度數據的關聯性分析結果進一步遴選出3組具專一性之管制重金屬代表性基因：鉻ｃｚｃＣ、鉻ｚｎｅＢ／ｈｍｘＢ及鋅ｃａｄＡ／ｙｖｇＷ，進一步設計共38組引子及探針進行ｑ－ＰＣＲ測試，測試結果從鉻的設計引子中選出Ｃｒ＿ｚｎｅＢｈｍｘＢ＿５以及從鋅的設計引子中選出Ｚｎ－ｃａｄＡｙｖｇＷ＿３１作為適用性最高的引子對。

中文關鍵字

第三代微生物總基因體定序、重金屬總基因體關聯性分析、分子檢測

類型	檔名	檔案大小	說明	最新版公開日期
期末報告	底泥品質快篩檢測技術研究-重金屬_成果報告（定稿本）.pdf	29MB		2024/12/26 上午 12:00:00

摘要	The pioneering “Tally and Design”molecular diagnostics development strategy de-ployed in this study involves first quantitating the distribution of microbial heavy metal resistant genes in contaminated sediment samples using Third Generation Metagenome Sequencing technology and customized bioinformatics pipeline. Next, the Meta-genome-associated analysis (MWAS) runs correlation analysis between the heavy metal resistant genes and concentrations of eight heavy metals to screen for most representative target genes. Lastly, homolog-based primer design will generate sequences to be used to synthesize q-PCR probe and test on sediment samples. Specifically, sediments samples were first collected from the irrigation canals of “Fan Ya Kou” and “Hsin Pi Chiu Tsun” in Changhua County, and subjected to metagenomic and heavy metal analysis respectively. The distributions of microbial heavy metal resistant gene in current and earlier studies were next calculated and used in downstream MWAS analysis. Consequently, czcC (Chromium), zneB/hmxB (Chromium) and cadA/yvgW (Zinc) genes were selected as the representative resistant genes of Chromiun and Zinc. Finally, a total of 38 sets of primer and probe were synthesized and validated, and Cr_zneBhmxB_5 and Zn_cadAyvgW_31were chosen as the most relevant probe and probe set.
英文關鍵字	Third Generation Metagenome Sequencing, Metagenome-wide association study with heavy metal, Molecular diagnostics

摘要

The pioneering “Tally and Design”molecular diagnostics development strategy de-ployed in this study involves first quantitating the distribution of microbial heavy metal resistant genes in contaminated sediment samples using Third Generation Metagenome Sequencing technology and customized bioinformatics pipeline. Next, the Meta-genome-associated analysis (MWAS) runs correlation analysis between the heavy metal resistant genes and concentrations of eight heavy metals to screen for most representative target genes. Lastly, homolog-based primer design will generate sequences to be used to synthesize q-PCR probe and test on sediment samples. Specifically, sediments samples were first collected from the irrigation canals of “Fan Ya Kou” and “Hsin Pi Chiu Tsun” in Changhua County, and subjected to metagenomic and heavy metal analysis respectively. The distributions of microbial heavy metal resistant gene in current and earlier studies were next calculated and used in downstream MWAS analysis. Consequently, czcC (Chromium), zneB/hmxB (Chromium) and cadA/yvgW (Zinc) genes were selected as the representative resistant genes of Chromiun and Zinc. Finally, a total of 38 sets of primer and probe were synthesized and validated, and Cr_zneBhmxB_5 and Zn_cadAyvgW_31were chosen as the most relevant probe and probe set.

英文關鍵字

Third Generation Metagenome Sequencing, Metagenome-wide association study with heavy metal, Molecular diagnostics