What are the steps involved in DNA based microarray?

There are five main steps involved in DNA based microarray. They are sample collection, isolation of mRNA, creation of labeled cDNA, hybridization, and collection and analysis. In the first step, 2 types of samples (healthy and infected cells) are generally collected for comparing and collecting the results. In step 2, the extraction of RNA from a sample is carried out by using a column or solvent such as phenol-chloroform. mRNA is isolated from the extracted RNA, leaving behind rRNA and tRNA. Since mRA has a poly-A-tail, column beads with poly-T tails are used to bind mRNA. After the extraction, a buffer is used to rinse the column to isolate mRNA from beads. In step 3, the reverse transcription of mRNA yields cDNA. Both the samples are then integrated with different fluorescent dyes for the creation of fluorescent cDNA strands. In step 4, the labeled cDNAs from both samples are placed on the DNA microarray which allows the hybridization of each cDNA to its complementary strand. They are then washed to remove unpaired sequences. In step 5, the microarray scanner is used to collect the data. The laser in the scanner excites the fluorescence of the cDNA, producing signals. The camera records images as the laser scans the array. Lastly, the computer inputs the data and yields results immediately. The distinct intensity of the colors for each spot determines the character of the particular gene.