[問題] TritonX-100 soluble and insoluble 的不同?
最近看 paper 的時候發現
有的人會分別收
TritonX-100 soluble and TritonX-100 insoluble cell extracts
來進行 Western blot
我想知道的是...
這兩種 fractions (TritonX-100 soluble and TritonX-100 insoluble)
所收集到的 proteins 性質上有什麼差異?
舉例來說:
我看到的這篇 paper 中,將細胞處理 MG132(a proteasome inhibitor)
原本他假設某個compound會促進某個protein進行proteasomal-deprendent degradation
所以預期加入proteasome inhibitor會block此target protein 進行degradation
但是他要看的 target protein 在 TritonX-100 soluble fraction 中並沒有增加
結果他又另外做了 TritonX-100 insoluble fraction 結果是增加的
於是作者提出結論認為此compound的確會促進
此 target protein進行proteasomal-deprendent degradation
所以我想知道TritonX-100 soluble 和 insoluble fraction 有何差異?
這是否代表 ubiquitinated proteins 是屬於 TritonX-100 insoluble?
我發現不少 paper 都會進行這樣的分離
但不是很清楚這樣做的目的何在?
有沒有人可以提供一些資訊呢?
補充說明:
For preparation of Triton X-100 soluble and insoluble fractions,
After removal of Triton X-100 soluble cell lysate supernatants
by centrifugation, the pellets were washed once with the lysis buffer
and 1x SDS loading buffer (50 μL) was then added to the pellets and
heated at 95°C for 15 minutes to dissolve the Triton X-100-insoluble proteins.
收cell lysates的lysis buffer含有低%的TritonX-100
本篇作者將離心過後的pellets再用SDS loading buffer回溶得到
Triton X-100 insoluble fractions
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◆ From: 61.216.103.208
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※ 編輯: patrickp1005 來自: 61.216.103.208 (11/13 20:25)
※ 編輯: patrickp1005 來自: 61.216.103.208 (11/13 20:32)
※ 編輯: patrickp1005 來自: 61.216.103.208 (11/13 20:33)
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