鑑定甲烷菌的寡核苷酸組、套組及彼等之鑑定方法Oligonucleotide set and kit for identifying methanogens, and method using the same

專利類型

發明

專利國別 (專利申請國家)

中華民國

專利申請案號

100134075

專利證號

I 456065

專利獲證名稱

鑑定甲烷菌的寡核苷酸組、套組及彼等之鑑定方法Oligonucleotide set and kit for identifying methanogens, and method using the same

專利所屬機關 (申請機關)

國立成功大學

獲證日期

2014/10/11

技術說明

本團隊成功地發展一項新型定性定量分子生物技術，應用於複雜環境樣本中的甲烷菌群進行高通量的分析。根據甲烷菌的親緣關係，本研究設計42個不同分類階層甲烷菌之專一性的核甘酸探針，經專一性、相容性、及靈敏性測試與最佳化實驗條件，成功配裝成8組的多通量分析探針套組，每套分析的通量數可達5-7個目標甲烷菌群。以建立的平台三重覆分析30個反應槽的厭氧污泥樣本，經22小時的分析後，完成2520 個偵測分析，可快速獲知這些樣本中偵測的6個目與33個屬甲烷菌群的相對豐富度數據，而且引子偵測專一性高，分析標準差小於12.2 %，靈敏度低達0.165 % or 107copies，顯示本技術可有效地進行反應槽污泥中甲烷菌群結構的定性定量分析，可快速提供改進反應槽操控所需甲烷菌群多樣性的指標資訊。本研究建立的高通量定性定量分生技術與分析策略可推廣應用於研究不同厭氧生物環境中甲烷菌的生態。 A newly developed molecular method was optimized to qualitatively and qualitatively analyze the methanogenic Archaea in the sludge from anaerobic reactors. In total, 42 oligonucleotide probes with specificity at the various hierarchical levels were designed and successfully applied in 8 multiplexing analyses. After optimization, the assay was used to detect the targeting methanogens in 30 different sludge samples, which 2520 detection reactions could be completed within 22 hrs, coming up with the abundance values of 6 orders and 33 genera methanogens with excellent detection specificity, reproducibility and sensitivity. The results showed that the detected methanogens could cover a total abundance of at least 79.3 % in the domain Archaea. Overall, the established high-throughput technology can effectively quantify the methanogens in engineered ecosystems and suggests that this platform developed in this study may be applicable for samples from other environments.

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