What is the workflow of illumina sequencing technology?

In a standard Illumina workflow, there are several key steps: preparing the library, amplifying clusters, sequencing DNA, analyzing images/base calling, discovering variants, and aligning reads. During library preparation, the DNA or RNA sample is processed to create a sequencing library. The library preparation involves fragmenting the DNA, adding sequencing adapters, and amplifying the fragments. The prepared library fragments are loaded onto a flow cell. This amplifies the clusters of DNA fragments, creating millions of clusters, each containing many copies of the same DNA fragment. Then, the amplified DNA or cDNA fragments, known as amplicons, undergo adapter ligation. This process attaches specific oligonucleotide sequences to the ends of the amplicons, permitting them to interact with a sequencing flow cell. During the sequencing step, each fragment of the library binds to primers and undergoes amplification, forming millions to billions of clonal clusters. Subsequently, fluorescently labeled nucleotides are used to build a complementary strand for each fragment. The sequencer then captures images of the flow cell, recording the emission from each cluster. By analyzing the fluorescent emission intensity and wavelength, the sequence of the templates is identified. The fluorescent signals are converted into base calls (A, T, G, or C). This process involves identifying the sequence of each cluster based on the emitted light. After base calling, the generated reads are aligned to a reference genome. This step involves mapping the reads to their corresponding positions on the reference genome, allowing for the identification of the location of each sequenced fragment. Variant discovery involves comparing the sequenced data to a reference sequence or other samples.