Resequencing is a method of genetic analysis that determines the DNA sequence of an organism for which a reference genome already exists. Its purpose is to compare an individual’s genetic code to this established standard. This comparative approach allows scientists to pinpoint differences and variations in the DNA. Identifying these genetic distinctions provides insights into the basis of traits, diseases, and evolutionary relationships.
The Resequencing Procedure Explained
The resequencing process begins with extracting DNA from a biological sample, such as blood, saliva, or tissue. This DNA is prepared for sequencing through library preparation, where long DNA strands are fragmented into shorter pieces. Special adapters are then attached to the ends of these fragments, allowing them to be recognized and processed by the sequencing machine.
The prepared DNA fragments are loaded into a Next-Generation Sequencing (NGS) instrument. These platforms read millions of these short DNA fragments simultaneously, generating vast amounts of sequence data in a relatively short time. The technology produces high-quality reads, typically less than 300 base pairs in length, providing a detailed snapshot of the individual’s genetic makeup.
A computational analysis follows the sequencing run. The newly generated sequence reads are aligned, or mapped, to the existing reference genome for that species. Computer programs compare each short read to the reference to identify its location. This alignment process highlights any discrepancies, which are flagged as potential genetic variations for further investigation.
Identifying Genetic Variations
Resequencing catalogs the differences that make each organism’s genome unique. One of the most common discoveries are Single Nucleotide Polymorphisms (SNPs). These are changes to a single “letter” in the DNA code and represent the most frequent type of genetic variation among individuals. Their prevalence makes them a focus for understanding genetic diversity and predisposition to certain conditions.
Another class of variations are insertions and deletions, collectively known as indels. These occur when a small number of DNA bases are either added to or removed from the genome. Their impact can be significant if they occur within a gene, potentially disrupting its function by altering how the genetic code is translated into proteins.
Resequencing also uncovers larger-scale structural variations like Copy Number Variations (CNVs). In CNVs, entire sections of DNA are duplicated or deleted, leading to an abnormal number of copies of a gene. Identifying these variations is how they can be linked to an individual’s traits, disease susceptibility, and response to medications.
Key Applications of Resequencing
In human health, resequencing helps understand the genetic basis of many conditions. In cancer genomics, scientists compare the DNA sequence of a tumor with a patient’s healthy tissue. This comparison helps identify specific mutations that drive the cancer’s growth, which can guide the selection of targeted therapies. It is also used to find genetic variants responsible for inherited disorders like cystic fibrosis or to assess risk for complex diseases.
Pharmacogenomics is a field that uses resequencing to study how a person’s genetic makeup influences their response to drugs. By identifying specific genetic variations, doctors can predict whether a medication will be effective or cause adverse side effects. This allows for a personalized approach to medicine, where drug choice and dosage can be tailored to the individual.
Resequencing is also used in population genetics to explore genetic diversity and trace human migration patterns. In agriculture, this technology identifies desirable traits in crops and livestock, such as disease resistance or higher yield, accelerating breeding programs. Conservation biologists also use it to assess the genetic health of endangered species to inform preservation strategies.
Resequencing Versus De Novo Sequencing
The choice between resequencing and de novo sequencing depends on whether a reference genome for the organism is available. Resequencing is used when a high-quality reference sequence already exists. Its goal is to detect variations like SNPs and indels by comparing the new sequence data against this established blueprint.
In contrast, de novo sequencing is used when studying an organism for the first time, where no reference genome is available. The term “de novo” means “from the beginning,” and this method involves assembling short sequence reads into a complete genome from scratch. This process is computationally intensive and aims to create the first reference genome for a species.
Consequently, resequencing is faster and more cost-effective than de novo sequencing because it bypasses the complex assembly step. The computational requirements are lower since the task is to align reads to an existing map rather than constructing a new one. A researcher would use resequencing to study genetic diversity in humans, whereas de novo sequencing would be necessary to map the genome of a newly discovered insect or plant.