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We further examined whether NHERF1 bound to PTEN at the endogenous expression level by using MCF7 and Zr75.1 cells that expressed high levels of NHERF1 and PTEN.
The expression of B5R and the replication of miRNA-regulated vaccinia virus (MRVV) with target sequences complementary to let-7a in the 3′-untranslated region (UTR) of the B5R gene depended on the endogenous expression level of let-7a in the infected cells.
All samples were run in triplicate and normalized by the endogenous expression level of GAPDH.
Nanomolar concentrations can be analyzed, compatible with protein studies at an endogenous expression level.
We first examined the endogenous expression level of SALL4 in SP cells in several hematological cell lines.
The endogenous expression level of these 4 miRNAs in LX2 and normal liver was low according to the microarray study (Figure S2).
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To disrupt Ago2 PICK1 interactions while maintaining endogenous expression levels of both proteins, we expressed Δ354 PICK1 to compete with endogenous PICK1 for Ago2 binding.
To investigate if TKT is abnormally expressed in oral cancer cells, we compared the endogenous expression levels of TKT among NHOKs, UM1 and UM2 cell lines.
We established ESC lines expressing FLAG-HA tagged version of WT and F266A WDR5 at near endogenous expression levels, and FPLC analysis confirmed that both tandem-tagged WDR5 proteins quantitatively formed equivalent MLL-WDR5 protein complexes with endogenous subunits.
We note, however, that the biological relevance of this phenomenon remains to be determined: it can occur when OFUT1 is over-expressed, but it is not clear whether it is significant at endogenous expression levels.
Unfortunately, the endogenous expression levels of SNAREs are often low which makes detection challenging.
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