NF-KB detection double-stranded DNA micro array chip and preparation

Abstract

The invention can prepare the microarray of double -stranded DNA(ds DNA) on the surface of solid substrates. By this method, the probes of ds DNA can engomphosised in the nuclear factor kappa B(NF-kappa B) to high-throughpput detect NF-kappa B inside of caryon and screen ds DNA / NF-kappa B molecules interfering dsDNA / NF-kappa B interaction. First, depending on patent technic and others methods of this invention, the invention prepare the microarray of double -stranded DNA(ds DNA) on the surface of solid substrates to allow mounts of dsDNA explorer on the dsDNA micro embattle to link with locus. Second, allow dsDNA micro embattle to across and link with specifically cell nuckoproteides or NF-kappa pure albumen. Third, do qualitative determination or quantiative determination to NF-kappa albumen in special caryon by reaction of NF-kappa B with dsDNA as micro embattle butt.

Description

1. Technical field [0001] In the field of basic molecular biology and biomedicine, the present invention proposes a new high-throughput detection technique for important transcription factor NF-κB, that is, NF-κB detection dsDNA microarray chip, and clarifies its various Preparation method: use the NF-κB detection dsDNA microarray chip prepared by various preparation methods proposed in the present invention to study the interaction law of transcription factor NF-κB and its DNA binding target in the study of gene expression regulation, high-throughput detection of cell nucleus In the fields of basic molecular biology and biomedicine, such as the expression and activation of various NF-κB in China, the screening of NF-κB DNA target binding drug small molecules, and NF-κB / DNA interaction interfering drug small molecules, etc. important application value. 2. Background technology [0002] Nuclear factor κB (Nuclear Factor kappaB, NF-κB) is a type of sequence-specific transcrip...

AI Technical Summary

Problems solved by technology

The advantage of this method is that it can use DNA microarray in situ synthesis technology to prepare high-density dsDNA microarray; but this method also has serious economic and technical problems, technically it relies on photolithographic mask in situ DNA microarray Synthetic patented technology, but the synthesis efficiency of single-stranded oligonucleotides on the surface of the solid phase substrate is not high, and the synthesis efficiency of each nucleotide is 92-96%. Only 4-20% of the oligonucleotides reach the required length of 40 bases. Therefore, the ssDNA and dsDNA microarrays prepared by photolithographic mask in situ synthesis technology are seriously polluted by truncated molecules.

Numerous competing truncated molecules would likely strongly inhibit and mislead protein binding experiments; and there is no reason to believe that every oligonucleotide on a microarray of single-stranded oligonucleotides synthesized in situ by a photolithographic mask could be primed oligonucleotides. Nucleotide binding, so that part of the 40-base full-length oligonucleotide cannot be converted to double-stranded nucleic acid, which not only further reduces the number of ideal double-stranded nucleic acid probes on the chip, but also those that are not converted to double-stranded Single-stranded oligonucleotides can also inhibit and mislead protein binding experiments

In addition, the dsDNA microarray prepared by this method is a bimolecular (bimolecular) dsDNA microarray, and there is a problem of probe stability, that is, the bimolecular dsDNA is affected by the experimental procedures such as binding and washing, and the double strands will be unzipped and lost. probe function; and this bimolecular dsDNA microarray can only be used once, so its usage efficiency is extremely low

Because the dsDNA microarray prepared by this method relies on the currently very expensive photolithographic mask in situ DNA microarray synthesis patent technology, coupled with the patent protection of this method itself, it is not only very expensive to apply, but also cannot be commercialized in my country. Develop application

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