The principle of DNA microarray technology is to integrate gene probes with known sequences onto a solid surface. A large number of labeled nucleic acid sequences from the tested biological cells or tissues are hybridized with the above probe array. By detecting the hybridized probes at the corresponding positions, rapid detection of genetic information is achieved.
Ⅰ. DNA Microarray Preparation Process
Chip preparation: DNA chip preparation methods include light-guided in-situ synthesis, chemical injection, contact spot printing, in-situ DNA control synthesis, non-contact micro-mechanical printing, and soft photolithography replication. Up to 400,000 different DNA molecules can now be placed on a 1 cm2 chip.
Sample preparation: This includes the isolation and purification of sample DNA or RNA, amplification of target gene fragments with PCR technology, and labeling of target genes.
Hybridization reaction: Suitable reaction conditions are selected to ensure that the reaction between biomolecules is under the optimal reaction conditions. Chip hybridization belongs to solid-liquid phase hybridization, and many factors affect hybridization, including target concentration, probe concentration, sequence composition of the hybridization partners, salt concentration, temperature, and washing conditions.
Detection and analysis of chip signals: The target genes in the sample specifically hybridize with the probes fixed on the chip and bind at different points on the chip. Fluorescent molecules are excited by specific wavelength light and emit specific wavelength fluorescence. The hybridized signal is obtained through a specific scanner. The chip scanners for chip scanning include laser confocal scanning chip and CCD chip scanner. The data obtained is processed using a dedicated processing system, including statistical and biological analysis of chip data, accumulation and management of chip databases, retrieval of chip-expressed genes on the international internet, and analysis of gene expression databases.
Ⅱ. The most prominent feature of DNA microarray technology is the ability to detect multiple samples at once
Different gene expression profiles of multiple genes have been successfully obtained by cDNA microarrays, which can detect the expression of more than 10,000 genes simultaneously. Therefore, DNA microarrays are a high-throughput and novel gene analysis method for parallel comparative analysis of multiple gene expression patterns in different materials. Compared with traditional methods for studying differential gene expression, it has the advantages of miniaturization, speed, accuracy, high sensitivity, and parallel detection of large amounts of information on the same chip.
DNA microarray technology has made significant progress in drawing gene expression profiles, finding target genes and functional genes, and other research areas. However, the disadvantage is that not all the sequences spotted are the ones needed for the experiment, and the required analytical instruments are relatively complex. In addition, DNA microarray technology is relatively limited in analyzing low-abundance transcripts. To ensure that a certain low-abundance transcript is included in the DNA microarray, a large number of clones need to be selected for amplification and spotting.
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