Detection of DNA binding protein with exonuclease protective DNA probe and hybrid DNA microarray chip

Abstract

The invention is to detect the DNA conjugated protein using the hybrid DNA micro-array slug of exonuclease to protect DNA probe. The process is: (a) preparing the combined protein with DNA probe, the vector PCR, Tag-DNA; (b) combining the combined protein with DNA probe and the DNA conjugated protein; (c) treating with the exonuclease; (d) PCR amplifying and marking; detecting the hybrid product of PCR and the Tag-DNA. The method can detect many kinds of the DNA conjugated protein simultaneously but not modifying and marking the protein.

Description

1. Technical field: [0001] DNA-binding protein is a kind of protein in proteomics that can combine with DNA to produce important functions. This kind of protein plays a role in gene expression regulation, DNA recombination , DNA repair (DNA reparation), DNA restriction (DNA restriction) and other cellular processes (cellular processes) play an important role. Some of these proteins exhibit DNA sequence-specific binding functions when interacting with DNA, such as various DNA-binding proteins and various restriction endonucleases that specifically bind to DNA. Because of the important role these proteins play in cellular processes, they have become important objects of functional genomics and proteomics research. Because this kind of protein is involved in the expression regulation of many genes, it is closely related to many diseases and becomes an important target (drug target) for diagnosis, treatment and drug research. The detection and analysis of DNA binding protein exp...

AI Technical Summary

Problems solved by technology

This method needs to label proteins with fluorescein in the detection, or prepare antibodies for proteins and label them with fluorescein to achieve chip detection. This not only requires experiments such as labeling proteins and preparing antibodies, but also the labeling may affect the DNA binding function of the protein.

This method is very effective when detecting the interaction between a protein and a large number of DNA on the chip; but it faces certain difficulties when detecting multiple proteins

It is particularly important that in practice, the objects to be detected are often nuclear extracts of tissues, cells, etc., and it is very difficult to fluorescently label such complex mixtures, especially for Those low-abundance DNA-binding proteins, it is very difficult to label them in nuclear extracts

In addition, if antibodies are used, it is necessary to prepare antibodies for all the proteins that need to be detected and label them with fluorescent light, so that they can be used for high-throughput detection of various DNA-binding proteins, which is obviously more difficult

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