产品用途

噬菌体表面展示系统辅助噬菌体

产品特性

产生用于测序和突变的单链噬菌粒DNA

产品简介

M13KO7是gII基因具有Met40IIe突变的M13 噬菌体。M13KO7具有P15A的复制起点和Tn903卡那霉素抗性基因，这两个片段插入至M13的复制起点。M13KO7在没有噬菌粒DNA的条件下也能复制。当存在一个携带有野生型的M13或f1复制起点的噬菌粒时，单链噬菌粒可以完整包装，并分泌到培养基中。该特性可用于生产用作突变和测序的单链噬菌粒DNA。为了繁殖和扩增M13KO7辅助噬菌体，需要使用TG1、ER2738或菌株XL1 Blue MRF′大肠杆菌菌株，因为这些菌株含有supE44突变，该突变提供谷氨酰胺抑制剂tRNA。M13K07辅助噬菌体严格用于从已切除的噬菌体中产生单链DNA（这也是VCSM13辅助噬菌体的用途）。

来源

M13KO7噬菌体上清是用标准程序从受侵染的E. coli ER2738中分离获得。

浓度

1.0 x 10¹¹ pfu/ml

规格

500ul/管，1管

贮存条件

1 x PBS和50%甘油

贮存温度

-20℃

扩增

1. Streak out TG1 onto an 2YT plate.

2. Pick a single colony of TG1 and inoculate into 5ml 2×YT and grow until OD600 = 0.3 (~2.5×10⁸ cells/ml).

3. Add helper phage at a multiplicity of infection (MOI) of 20:1 (phage-to-cells ratio).

Note: If amplifying M13K07 helper phage, add Kanamycin to a final concentration of 25μg/ml to the medium 30 minutes after the helper phage and cells have been allowed to grow together.

4. Grow the culture at 37℃ with vigorous aeration (~300rpm) for 8 hours.

5. Heat the culture to 65℃ for 15 minutes.

6. Spin down the cell debris and transfer the supernatant to a fresh tube.

7. Titer the helper phage produced.