1ml GST Syringe Kit

————依靠注射器加压或重力分离两用预装柱，

■包装内容

GST Syringe 层析柱（预装1 ml GST Sepharose）

10支

10x STE Buffer (pH 8.0)                        50ml

10x PBS (pH 7.4)                              50ml

10% Triton X-100 in STE                      50ml

GSH                                          0.5g

■      产品特点

■ 产品说明：

GST标签纯化树脂能简单快速地纯化谷胱甘肽-S转移酶（GST）、谷胱甘肽依赖性蛋白和谷胱甘肽-S转移酶重组衍生物，尤其适用于在大肠杆菌、昆虫细胞和哺乳动物细胞中表达的GST融合蛋白。高亲和力GST标签蛋白纯化树脂可直接将GST融合蛋白从细菌溶解物中纯化出来，且具有极好的结合能力，每毫升GST纯化介质可结合多于10 mg的GST融合

蛋白蛋白纯化流程如下：

1ml GST Syringe Kit

实验操作方法

1.Seed 5 ml of overnight culture to 500 ml LB with 150 m Induce with 0.1 mM to 2 mM of IPTG. Grow for 3 hr at 37℃ or grow overnight at room temperature.

Lower IPTG concentrations and lower growing temperatures tend to produce greater solubility at the expense of yield.

4. Pellet cells by centrifuging at 3000g 4℃for 10 min. Decant media and resuspend cells in 30 ml ice-cold PBS to wash. Transfer to a 50ml tube and centrifuge at 3000 g, 4℃for 10 min. Decant PBS.

5. This is a convenient point to stop and to store pellets at -80℃ or -20℃.

6. Thaw pellet on ice if cells are frozen else proceed to the next step.

7. Resuspend pellet in 40 ml of ice cold STE Buffer.

8. Add lyozyme to final concentration 1mg/ml, incubate on ice for 15 min. Just before sonication, add DTT o final concentration 1Mm and Mix thoroughly by inversion and sonicate for a total time of 10 min (or a appropriate time),still to the lysate becomes transparent.

9. Centrifuge 12,000 rpm for 20 min. Transfer supernatant to a 50ml conical tube and discard the pellet. Add 10 ml of 10% Triton X-100 toThe effective concentration of Triton X-100 will be 2%. Incubate at room temperature for 30 min.

10. Wash the prepared Glutathione Sepharose with 10 ml PBS,

11. Pour the lysate to 1 ml Glutathione Sepharose column with a flow of 2-3ml/min.

12. Wash the prepared Glutathione Sepharose with 10 ml PBS T

13. If desired, elute with 10 x 1 ml fractions of Elution Buffer (STE with 20Mm GSH) . Determine desired fractions with SDS PAGE.