DNA mutation and gene editing concept

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DP Biology

Mutation and Gene Editing

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Introduction What Are Mutations? Types of Mutations Causes of Mutations Effects of Mutations Detecting Mutations Gene Editing CRISPR-Cas9 Base Editing (BE) Prime Editing (PE) Gene Therapy Ethical Considerations Summary

Mutation and Gene Editing

DNA is the blueprint of all the living things on this planet, including us, and it is passed on from one generation to another during reproduction. DNA is responsible for defining the characteristics of living things and controlling the functioning of the cells.

Mutations, or changes in the DNA sequence of a gene, can occur in a single nucleotide of a gene or in a large section of a chromosome. A mutation can be harmless or harmful. There are mutations that can benefit the organism.

Biologists study mutations to understand biological processes. Knowledge of mutations assists in the understanding of diseases, the improvement of crops, and the discovery of new therapies.

What Are Mutations?

Biologists understand changes in the DNA sequence of a gene to be mutations. There are internal and external factors that can cause mutations in an organism. These changes can be permanent, and if the mutation occurs in the gametes, the mutation can be passed to the next generation.

Mutations can occur:

  • Spontaneously: due to errors during DNA replication.
  • Induced: mutations can be caused by exposure to chemicals, radiation, or viruses.

Types of Mutations

There are many ways to categorize mutations, but here are some common examples.

Point Mutations

A point mutation involves a change in a single DNA base pair. There are three main types of point mutations:

Silent mutation
A nucleotide is changed, but the resulting amino acid is unchanged. There is no effect on the functioning of the protein.
Missense mutation
The amino acid affected by the nucleotide change is altered, thus the protein may function differently or not as effectively.
Nonsense mutation
A nucleotide change results in the creation of a stop codon. This causes protein synthesis to halt earlier than normal, usually resulting in a protein that is non-functional.

Frameshift Mutations

These mutations occur when one or more nucleotides are added or lost in the DNA sequence. This causes the entire reading frame of the gene to shift.

Example: If one nucleotide is deleted, the amino acids that come after this deletion are changed.

These types of mutations are typically the most harmful, as they can alter the function of the entire protein.

Chromosomal Mutations

These types of mutations impact larger structures in the chromosomes, often including many genes:

Deletion: a section of a chromosome is lost.
Duplication: a section is repeated.
Inversion: a section is reversed.
Translocation: a section of one chromosome moves to another chromosome.

Causes of Mutations

There are a number of both internal and external reasons as to why a mutation may occur.

Internal Causes (Spontaneous)

  • During DNA replication, DNA polymerase can randomly insert an incorrect nucleotide. This is considered an error and is a reason for a spontaneous mutation.
  • Deamination and depurination are types of spontaneous chemical changes.

External Causes (Induced)

  • Chemicals – Chemical mutagens like benzene and mustard gas directly change DNA bases.
  • Physical – Ultraviolet light, X-ray, and gamma-ray interaction with DNA.
  • Biological – Viruses that insert their own DNA into a host organism.

Effects of Mutations

The effects of a mutation can be classified into three categories:

Useful
A mutation in the CCR5 gene can result in a person being unable to contract HIV.
Neutral
Survival or reproduction is not affected by the mutation.
Harmful
The mutation can lead to decreased fitness or genetic disorders like cystic fibrosis or sickle-cell anemia.

Detecting Mutations

Detecting the presence of a mutation can be done using several modern techniques in molecular biology:

  • PCR (Polymerase Chain Reaction) – This technique amplifies a section of DNA so that changes can be seen.
  • Gel electrophoresis – Used to separate DNA fragments so that the insertion of a DNA fragment or deletion can be detected.
  • DNA sequencing – In figuring out the presence of point mutations, DNA sequencing is useful in that it provides the exact sequence of nucleotides.

Gene Editing

Gene editing is the modern and more precise technique of altering DNA sequences to either study the DNA or to enhance certain DNA traits. This gene editing technique is a lot more precise and faster than traditional techniques like selective breeding.

CRISPR-Cas9

CRISPR-Cas9 is a game-changing gene-editing technique in that it allows the cutting of pre-determined locations on DNA so that certain DNA sequences can be added, removed, or changed. This technique has its origins in the bacterial defense system.

CRISPR is made up of several parts:

  • gRNA is programmed to bind to a specific part of a target DNA sequence.
  • The Cas9 enzyme is used to 'cut' the part of the DNA that the gRNA is bound to. It is a region of double-stranded DNA.

Repairing the break in DNA is done via the cell's DNA repair mechanisms. Two different cellular DNA repair mechanisms will repair the breaks:

  • Non-homologous end joining (NHEJ): When repair is done via this mechanism, it will yield small random insertions or deletions. This mechanism is often used to knock out or deactivate a specific gene.
  • Homology-directed repair (HDR): This repair mechanism utilizes a template strand of DNA that is the sample DNA that will be inserted.

CRISPR can be used/ is used to:

  • Correcting genetic disorders in lab cells.
  • Getting crops to not get sick (developing disease-resistant crops).
  • Finding out how genes work in model organisms.

Base Editing (BE)

Base Editing (BE) is a more refined version of the gene editing technologies, and in fact, Base Editing is more precise than CRISPR. Instead of cutting DNA, it converts one nucleotide base into another.

Example: To change a 'C' to a 'T' (without breaking the DNA strand) is possible with base editing. This improves precision in editing and reduces the unwanted modifications that are often seen with CRISPR. It is useful for point mutations (changes in DNA sequence that correspond to a single nucleotide base pair) that cause ill health/disease, and for correcting genetic disorders.

Prime Editing (PE)

Prime Editing (PE) is a newer and more advanced gene editing technology that is currently available. It is used to insert or delete or both, change the sequence of DNA, and do so without causing double-strand breaks.

This is done using a specific type of guide RNA known as the prime editing guide RNA (pegRNA) to specify which part of DNA is to be changed.

Incredible as it may be, the Top End Guide (TEG) RNA and the Reverse Transcriptase (RT) enzyme are used to write a new sequence of DNA.

In comparison to CRISPR, this technology is more precise and less damaging than CRISPR and is therefore better to use for gene therapy.

Gene Therapy

Gene therapy is a technique that treats genetic diseases by putting healthy genes into a patient's cells.

Gene therapy can be somatic, meaning it's a body cell change that will not be passed down, or germline, which alters egg/sperm genes and can be passed down.

Some of the methods gene therapy can be done by include:

  • Viral vectors, where the genes are delivered using a modified virus.
  • Non-viral methods, which include things like lipid nanoparticles or electroporation (the application of a localized electrical field to stimulate cells to allow the passage of ions and molecules).

Example: The addition of a functional gene into a patient's immune cells in the treatment of severe combined immunodeficiency (SCID).

Ethical Considerations

Gene editing brings a lot of ethical questions, including:

  • Should we allow germline editing, which will affect the lives of generations in the future?
  • The chance of having unintended or off-target mutations.
  • If wealth becomes a factor, will it just be the wealthy people or the wealthy countries that will be able to use these treatments?

Scientists have to be very cautious in following the restrictions, particularly where the edits could be passed down.

Summary

  • Mutations are changes that occur in DNA. There are different types of mutations, like point mutations, frameshift mutations, or chromosomal mutations.
  • Mutations can occur due to several things, such as spontaneous oversight, exposure to chemicals, radiation, and some viruses. The effects of mutations can be an improvement, remain the same, or be detrimental.
  • To find mutations, we can use PCR (Polymerase Chain Reaction), electrophoresis (the movement of charged particles in a fluid or gel), and DNA sequencing.
  • Gene editing is a way to change DNA in a precise manner. It can use CRISPR, which is a method of genetic engineering that involves the modification of DNA, and base and prime editing (which are also methods in genetic engineering).
  • Gene therapy means that we can treat genetic diseases. This can be done using methods that are viral or non-viral in nature.
  • When we edit genes (especially in humans), we have to be really careful since there are ethical issues involved.

The rapid nature of DNA can be seen through the lens of mutation and gene editing. While mutations can be seen as a driving force of evolution, gene editing can be seen as humanity's first attempt to improve or alter DNA intentionally. Both concepts are foundational to understanding life at a molecular level. Furthermore, both concepts serve as a basis for the further advancement of medicine, agriculture, and biotechnology.