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• W2068047648 abstract "RAP30, an RNA polymerase-associated protein (RAP) of approximately 30 kDa, is a component of the eukaryotic general transcription factor IIF (TFIIF). We have isolated a monoclonal antibody (MAb) that can be used to purify RAP30 under nondenaturing conditions. This MAb (designated 1RAP1) is a unique type of MAb that we have designated “polyol-responsive MAb.” Polyol-responsive MAbs are high-affinity antibodies that release antigen in a buffer containing a low-molecular-weight polyhydroxylated compound (polyol) and a nonchaotropic salt. RAP30, contained on pET11d, was expressed in Escherichia coli by culturing and inducing protein expression at 26°C. Under these conditions, approximately 50% of the RAP30 remains soluble. Inclusion bodies were removed from the cell lysate by centrifugation, the supernatant was treated with polyethyleneimine at 0.5 M NaCl to remove nucleic acids, and the soluble protein was applied directly to MAb-conjugated Sepharose. After extensive washing, RAP30 was eluted with buffer containing 0.75 M ammonium sulfate and 40% propylene glycol. RAP30 produced by this procedure stimulates transcription from a minimal promoter. This is a rapid method for purifying unmodified RAP30 without renaturing the protein from inclusion bodies." @default.
• W2068047648 created "2016-06-24" @default.
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• W2068047648 date "1999-11-01" @default.
• W2068047648 modified "2023-09-25" @default.
• W2068047648 title "Immunoaffinity Purification of the RAP30 Subunit of Human Transcription Factor IIF" @default.
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• W2068047648 doi "https://doi.org/10.1006/prep.1999.1128" @default.
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