[求救] 純化soluble protein時~在binding buffer可添加TitonX100?
請問各位 如果我想要純化soluble protein
在誘導後離心去掉上清液後
以binding buffer回溶(之後要用 Ni-NTA column作純化)此時的binding buffer
是50mM tris-HCl
500mM NaCl(可以不加這個嗎 是不是可以更容易在wash的時候洗掉非專一性的
蛋白質呢)
2mM PMSF
可以加0.5-1%的Triton X-100嗎?會不會造成他的構型改變呢?
此時的imidazole我查到的資料有兩種說法耶
一種是binding buffer不含imidazole resin的結合比較好 比較不會bind到雜蛋白
一種是大約5-10mMimidazole 說是這樣比較可以先洗掉一些非專一性的雜蛋白
以上問題 麻煩大家><感謝
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