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題名 | 凍熱細菌質體轉形法=Freezing-warming Method for Transformation of E. coli |
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作者姓名(中文) | 王強生; 廖仁盟; | 書刊名 | 中華農業研究 |
卷期 | 48:1 1999.03[民88.03] |
頁次 | 頁60-63 |
分類號 | 430.16341 |
關鍵詞 | 細菌轉形法; 凍熱法; 大腸桿菌; Transformation; Freezing-warming method; E. coli; |
語文 | 中文(Chinese) |
中文摘要 | 細菌轉形為分子生物學相關研究領域實驗室內操作最普遍之實驗步 驟。本文介紹我們所發展之一快速且簡單之細菌轉形法名為「凍熱細菌質體轉形 法」,利用此一方法可快速且大量進行細菌之轉移,轉形效率可達8.7x106轉形 株/μg質體DNA。此一凍熱處理細菌質體轉形法僅須15min,較現有大多數方法 耗時75-90min可節省60-75min,顯著縮短轉形所需時間。本凍熱處理細菌質體 轉形法其步驟如下:將質體DNA與100 μl之細菌勝任細胞混合後,置於液態氮 內5分鐘進行凍處理,再立即移至37℃進行熱處理5分鐘後,直接塗佈於已在 37℃回溫之選擇性培養基,雖有助於提高細菌轉形效率,但較為費時。本試驗結 果顯示,此一凍熱處理細菌質體轉形法,具快速(少於15分鐘)、簡單、便宜及 可大量操作等優點,適合任何分子生物實驗室之經常性使用。 |
英文摘要 | We report a freezing-warming method for transformation of E. coli. In this method, plasmid DNA were mixed with 100 μl ice cold competent cells, freezing treated in liquid nitrogen for 5 min followed with heat treatment at 37℃ for 5 min, then directly spread onto the selective agar plate pre-warmed to 37℃ and cultured for 16-18 hr. Addition of recovery culture at 37℃ for 30 min before plating increased transformation efficiency several folds, however more time was required. The advantage of this new transformation method is time-saving. In gerneral, a transformation efficiency up to 8.7 × 10[8fb6] colony forming units/μg DNA can be achieved by this method within 15 min and it is significantly faster than most of the reported methods that usually require 75-90 min. |
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