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頁籤選單縮合
題名 | 偵測脫氮性Pseudomonas Sp.之DNA探針製作研究=Studies on Making the DNA Probe to Detect the Denitrifier Pseudomonas Sp. |
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作者 | 鄧德豐; 魯祖平; Dent, Der-feng; Lu, Tzu-ping; |
期刊 | 國立屏東科技大學學報 |
出版日期 | 19980900 |
卷期 | 7:3 1998.09[民87.09] |
頁次 | 頁187-193 |
分類號 | 432.225 |
語文 | chi |
關鍵詞 | 聚合酵素鏈鎖反應; DNA聚合酵素; 模板DNA; 引子; PCR; DNA polymerase; Template DNA; Primer; |
中文摘要 | 以人工合成364-368 (5'∼3' : GAC GTG GTG GTC GGC)之DNA片斷與582-586 (5' ∼3' : GTG GTT GAC CAT GAC)之DNA片斷,418-422 (5'∼3' : AGG CAG AAG CTT GAC) 之DNA片斷與582-586 (5'∼3' : GTG GTT GAC CAT GAC)之DNA片斷,以及528-532 (5'∼ 3' : GAG GAG GAC AAC AAG)之DNA片斷與582-586 (5'∼3' : GTG GTT GAC CAT GAC)之 DNA片斷等三組來當引子(primer),與另外從兼具硝化與脫氮性之P. aeruginosa抽出之 genomic DNA當模板(template)DNA,在下列條件下〔變性(denature)95℃、10sec,融接 (annealing)60℃、10sec,聚合反應(polymerization)72℃、20sec,50循環(cycle),S:8.0〕進行 聚合酵素鏈鎖反應(polymerase chain recation)合成探針,發現在聚合酵素鏈鎖反應時其最適 反應系為:Mg□濃度在20mM以上,dNTP在1.0-2.0mM,Taq polymerase在757.4 μg/μl 以上,引子濃度在50pM以上,融接溫度在60℃,較高的循環數可以比較安定的合成654bp 及492bp二條特異性DNA片斷,162 bp的DNA片斷之合成因較短故其再顯性較差。654 bp 及492bp二條DNA片斷與具有硝化及脫氮性P. aeruginosa均具有專一性的反應,對其他供 試菌均不會反應,故此二條DNA片斷可做為偵測硝化與脫氮性P. aeruginosa之良好的探針。 |
英文摘要 | The artificial proimers of 364-368 (5'∼3' : GAC GTG GTG GTC GGC) and 582-586 (5' ∼3' : GTG GTT GAC CAT GAC) DAN Fragment, 418-422 (5'∼3' : AGG CAG AAG CTT GAC) and 582-586 (5'∼3' : GTG GTT GAC CAT GAC) DNA frgment, and 528-532 (5'∼3' : GAG GAG GAC AAC AAG) and 582-586 (5'∼3' : GTG GTT GAC CAT GAC) DNA fragments were synthesized, respectively. Then these primers were used in PCR with the template DNA extracted genomic DNA from both nitrifying and denitrifying bacteria of P. aeruginosa, under the condition of Denature)95℃、10sec, Annealing 60℃、10sec, Ppolymerization 72℃、20sec, 50 cycle, S:8.0. The optimum reaction system for PCR was found at Mg□ above 20mM, dNTP at 1.0-2.0mM, Taq polymerase above 757.4 ug/ul, primer concentration above 50pM, annealing temperature at 60℃, and higher cycle numbers were more stable for synthesizing the specific DNA fragment of 654 bp and 492 bp. The shorter DNA fragment of 162 bp was lesser reappearance in PCR. Two DNA fragment of 654 bp and 492 bp were specific reacted with the nitrifying and denitrifying bacteria of P. aeruginosa, there were no reaction with other test micor-organisms, so, this two DNA fragment were used as the best probe for detection the nitrifying and denitrifying P. aeruginosa. |
本系統之摘要資訊系依該期刊論文摘要之資訊為主。