甘藍基因轉移之研究（一）基因鎗轉移法

Studies on Gene Transfer Into Cabbage (Brassica oleracea L. var. capitata) (I) Airgun Transfer

本研究以甘藍為材料，利用基因鎗轉移的方法，將GUS報導基因轉移到甘藍，並以轉殖後之GUS基因表現的活性為依據，探討轉移之適當條件。試驗結果顯示，基因鎗轉移GUS基因到甘藍原生質體，以氣室真空度在200 mmHg，2.25 X 10⁵原生質體（150μl）為最適宜。轉移至甘藍組織以胚及莖頂分生組織為材料較佳，設定氣室真空度為600 mmHg、鎗口至檔板間距離為4公分，取5至lOμ（lμg/μl）的環狀DNA，扣板機一次，鎗擊五個胚，可得到最佳的轉移效果。基因鎗轉移不同質體的效率，以基因較小者較佳。利用基因鎗轉移基因到甘藍成熟胚，是一種簡便、快速分析基因暫時表現的方法。但轉殖的胚會隨組織發育逐漸消失GUS活性，顯示GUS基因並非穩定嵌入甘藍的染色體。

The objective of this study is to establish an optimal condition of airgun transformation system for cabbage (Brassica oleracea L. var. capitals) by GUS reporter gene. Results indicated that setting the chamber pressure at 200 mmHg, and applied 150 μ 1 of cabbage protoplasts (2.25 X 10⁵) was the best condition for airgun transformation. The highest level (100%) of transient GUS gene expression were observed both in the mature embryo and shoot meristera when the transformations were conducted at the following conditions, i. e., the distance between stopping plate and material is set at 4 cm, chamber vacuum is at 600 mmHg and using circular plasmids. The expression of GUS activity was decreased drastically since 6 days after transformation indicated that the expression of GUS gene was not due to the stable integration but only a transient gene expression.