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以流式細胞儀判讀人為誘發鯉魚三倍體之探討

  • 出版日期:82-12-01
  • 標題title(英):
    Investigation on Artificially Induced Triploidy in Common Carp Using Flow Cytometer
  • 作者:趙乃賢, 劉富光, 黃家富, 楊志斌, 廖一久
  • 作者auther(英):Chao, N. H., F. G. Liu, C. F. Huang, C. P. Yang, I
  • 卷別:1
  • 期別:2
  • 頁碼:39-54

鯉魚 (Cyprinus carpio) 經由各種方式(如冷擊、高水壓、化學處理)誘發之後 ,必需經由染色體套數或 DNA 含量和對照組之比對來確認其三倍體率。本實驗室採用 的方法有一、染色體圖法 (Karyotyping method);二、銀染法(Silver staining method);三、粒度分析儀法(Coulter counter method);四、流式細胞儀法 (Flow cytometer method)。前三種方法本實驗室已陸續引進使用並不斷改進中,目前則嘗試 使用流式細胞儀作鑑定的工作。此期間不僅從結果分析來推論各種三倍體誘發方法的可 行性及其成效,而且和曾經採用之其他方法比較優缺點。
流式細胞儀分析法經多方測試,確知其特點為直接測知和 DNA 專用螢光染劑結合 之細胞核染色體內之 DNA 含量,而且藉助自動分析之功能,能正確且快速判讀染色體 倍數性,值得提供經常且大量測試魚類多倍體之用。有關誘發之結果顯示以 1℃ 冷擊 處理在不同組得到 66.6、80.4、86.2 及 86.4% 之三倍體鯉魚,而高水壓和高鈣高pH 化學處理組三倍體率則偏低,甚或為零。

摘要abstract(英)


After the induction of triploidy in carp (Cyprinus carpio) by cold shock, chemical shock, or hydrostatic shock, the identification of triploidy presence using flow cytometer to determine DNA content was developed and compared with three other methods, including karyotyping method, silver staining method and Coulter counter method, which were previously used in our laboratory. Three major protocols A, B and C adopted for pretreatment of red blood cells were found to be feasible and pratical with stepwise improvement of easy manipulation. The treated RBC samples were run in a flow cytometer allowing the sample injection at optimum flow rate (15~30 *l/min) sheath pressure (7.51~15.00 PSI) and laser power (0.15 MV and 8.62 A) to obtain better half pick coefficient of variation (< 5%) and histograms of fluorescence intensity from RBC stained with PI (Propidium Iodide). The histograms were analyzed using the supplied DNA analysis program and a recommended software. The results indicated that the flow cytometer has the advantages of being accurate and capable of checking more than 10,000 cells within 3 min. In this study, cold shock of 1℃ resulted in triploidy of 66.6, 80.4, 86.2 and 86.4% in various experimental groups, while high Ca, high pH solution treatment and hydrostatic pressure shock gave poor production of triploidy.