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DNA Microarray: Principle, Types, and Steps Involved
DNA microarrays are powerful tools for analysing gene expression patterns, detecting DNA mutations, and identifying genetic variations. They are used extensively in research, clinical diagnosis, and drug discovery. The microarray technology is based on the hybridization of labelled DNA or RNA molecules to complementary sequences that are immobilized on a solid substrate. In this article, we will discuss the principle, types, and steps involved in the DNA microarray technology.
Principle of DNA Microarray
The principle of DNA microarray technology is based on the hybridization of complementary sequences of nucleic acid molecules. The technology involves the preparation of a microarray chip, which consists of a solid substrate, usually glass or silicon, with thousands or millions of immobilized DNA probes. The probes are complementary to specific sequences of DNA or RNA molecules that are to be analysed. The DNA or RNA molecules that are labelled with fluorescent dyes are hybridized to the probes on the microarray chip. The intensity of fluorescence is measured to quantify the amount of hybridization that has occurred between the probes and the target molecules.
Types of DNA Microarray
There are two main types of DNA microarrays: cDNA microarrays and oligonucleotide microarrays.
cDNA microarrays are prepared by spotting cDNA fragments on a microarray chip. The cDNA fragments are prepared by reverse-transcribing mRNA molecules into cDNA using reverse transcriptase. The cDNA fragments are then labelled with fluorescent dyes and hybridized to the probes on the microarray chip.
Oligonucleotide microarrays are prepared by synthesizing oligonucleotides directly on the microarray chip. The oligonucleotides are designed to be complementary to specific DNA or RNA sequences that are to be analysed. The target molecules are labelled with fluorescent dyes and hybridized to the probes on the microarray chip.
Steps Involved in DNA Microarray
DNA microarray technology involves several steps, including sample preparation, microarray fabrication, hybridization, washing, scanning, and data analysis.
The first step in DNA microarray analysis is sample preparation. The sample can be RNA, DNA, or protein, depending on the purpose of the analysis. The sample is extracted from cells or tissues using various methods and then purified to remove impurities and contaminants.
The next step in DNA microarray analysis is microarray fabrication. The microarray chip is prepared by depositing DNA probes onto a solid substrate, usually a glass slide or silicon wafer. The probes can be cDNA fragments or oligonucleotides, depending on the type of microarray.
The third step in DNA microarray analysis is hybridization. The sample is labeled with fluorescent dyes, either directly or indirectly, and then hybridized to the probes on the microarray chip. The hybridization occurs between the complementary sequences of the probes and the target molecules.
The fourth step in DNA microarray analysis is washing. The microarray chip is washed to remove unbound or non-specifically bound molecules. The washing conditions are carefully optimized to minimize nonspecific binding and maximize specific binding.
The fifth step in DNA microarray analysis is scanning. The microarray chip is scanned using a fluorescence scanner to measure the intensity of fluorescence at each spot on the chip. The intensity of fluorescence is proportional to the amount of hybridization that has occurred between the probes and the target molecules.
The final step in DNA microarray analysis is data analysis. The data obtained from the microarray chip are analysed using various software tools to identify differentially expressed genes, detect DNA mutations, and identify genetic variations.
Q1. What is a DNA microarray?
Ans. A DNA microarray is a tool used to measure the expression levels of thousands of genes simultaneously. It consists of a small glass slide or silicon chip with thousands of small spots, each containing a different DNA sequence that represents a single gene.
Q2. What is the principle of DNA microarray?
Ans. The principle of DNA microarray is based on the hybridization of DNA strands. The DNA microarray contains thousands of known DNA sequences that are complementary to the DNA of the sample being tested. The DNA from the sample binds to the complementary sequences on the microarray, and the amount of bound DNA is measured to determine the expression level of each gene in the sample.
Q3. What are the types of DNA microarray?
Ans. The two main types of DNA microarray are the cDNA microarray and the oligonucleotide microarray. The cDNA microarray uses complementary DNA as probes, while the oligonucleotide microarray uses short DNA sequences as probes.
Q4. What are the applications of DNA microarray?
Ans. DNA microarray has many applications, including −
Gene expression profiling.
Identification of genetic mutations.
Diagnosis of diseases.
Agriculture and biotechnology research.
Q5. What are the advantages of DNA microarray?
Ans. The advantages of DNA microarray include −
High throughput: It can analyse thousands of genes at once.
Accuracy: It can detect small changes in gene expression levels.
Sensitivity: It can detect low abundance transcripts.
Reproducibility: It can produce consistent results.
Cost-effective: It can analyse multiple samples simultaneously.
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