Method for enriching and separating endogenous transcription factors and compounds thereof and concatenated transcription factor response elements special for method

Abstract

The invention discloses a method for enriching and separating endogenous transcription factors and compounds thereof and concatenated transcription factor response elements special for the method. The method includes of designing multi-copy double-stranded DNA (deoxyribonucleic acid) binding elements according to the characteristic that transcription factors are bound with sequence-specific DNA elements, adding double-stranded DNA arms labeled with biotin to two ends of each DNA binding element by the aid of molecular cloning technology to form 'DNA baits', coupling the biotinylated DNA baits which are purified onto magnetic beads enveloped by streptavidin when in separation and purification of the endogenous transcription factors and compounds thereof, incubating the magnetic beads together with the prepared nucleoprotein, and finally separating and purifying the endogenous transcription factors and the compounds thereof from the nucleoprotein. Further, after the endogenous transcription factors and the compounds thereof are separated and purified, protein identification and function analysis of the transcription factors can be performed subsequently.

Description

technical field [0001] The invention belongs to a method for enriching and separating target proteins in the field of biotechnology, and in particular relates to a method for enriching and separating endogenous transcription factors and their complexes and a special transcription factor tandem binding sequence. Background technique [0002] About 6% of the genes in the human genome encode transcription factors, the second largest class of proteins encoded by the human genome. Transcription factors not only play an important role in the regulation of gene expression, but also are key nodes in the intracellular signal transduction network. Various signaling pathways for cells to sense intracellular and extracellular stimuli are cross-linked by transcription factors, thus forming a complex signal network. Therefore, research on transcription factors and their complexes has been receiving great attention. However, the expression abundance of transcription factors in cells is e...

AI Technical Summary

Problems solved by technology

[0004] The expression levels of transcription factors are often very low, and it is often difficult to analyze transcription factor expression profiles at the proteomic level using traditional methods

When analyzing the expression profile of transcription factors or their dynamic changes, specific reagents (such as antibodies) and immunoprecipitation strategies are required to enrich and separate transcription factors during the sample preparation process. However, due to the relatively high cost of antibodies, the types of antibodies Limited, the application of immunoprecipitation with antibodies to enrich and isolate transcription factors is greatly limited, and this strategy can only analyze individual transcription factors, making it difficult to achieve large-scale enrichment, isolation and identification of all transcription factors

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