g418如何配制
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问:
最近要做稳定转染了,但不知道如何配制G418,有一些细节想问问大家,我打算用PBS来配,我想问问是用高压过的PBS来配,还是用未高压的PBS配,配完了是不是要过滤,然后放到-20保存,用的时候是不是放在4度.请各位来帮我解答一下,谢谢.
答:
我实验室的师兄一直是用HEPES配制的,因为PBS对细胞的损伤会大一些。
配的时候就是用配好的HEPES溶G418,然后过滤,不用高压灭菌了,配好后分装,-20度保存,用的时候取一管放4度。
100mM HEPES(pH7.3)缓冲液(500ml)
ddH2O 490ml
Nacl 4g
Na2HPO4-12H2O 0.1357g
Kcl 0.185g
Glu 0.5g
HEPES 2.5g
用氢氧化钠调PH至7.3 ,定容至500ml
G418贮存液(50mg/ml)的配制
G418(merck分装),效价大于等于700U/mg
按实际效价配制G418 solution.1g G418粉加14ml 100mM HEPES(pH7.3)溶解
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