Gene editing is trending these days. A revolution in Biology, that has all the potential to cure the suffering. All thanks to the technology, that led to this magnum opus in the field of Genomics. The birth of Genomics was laid down in the experiments of Mendel.
The scientific community today is well aware of Mendel, the father of Genetics, who called the hereditary units as cell elements. After 25 years of his death, Wilhelm Johansen, in 1909 named the hereditary unit with a catchier word, the gene.
From then on, genes have become the fundamental concept in biological sciences and caught the interest of many researchers. Discoveries unravelled many fascinating details about genes, which slowly extended the development of genetics into an offshoot in the field of biology, Genomics.
Genomics and Genetics are usually confused. Genetics is all about the heredity of genes and their variation. Genomics is the study of genes.
Important periods and events in the development of Genomics:
Genomics started with DNA sequencing in 1970, whose data shed light on the structure and functioning of a gene. The years before 1970 saw the discovery, genetic nature, composition, structure of DNA and also the central dogma of life.
After the 1970s, technological advancements have driven genomics to its present status. The techniques like Polymerase chain reaction (PCR) and Gene sequencing, have progressed the deciphering of the entire genome of microbes. These events have set the stage for the Mapping and Sequencing of the entire Human Genome.
The status of Genomics is further fuelled to newer miles with the automation and computing power of supercomputers. From then on, there has been a data explosion on the genetic information available, which is being stored securely in databases. This information helps us study biological problems at a level never possible before.
The following are the key areas, where Genomics will have a major impact:
The above points proclaim the importance of Genomics in the field of Biology. Genomics Assignment Help provides a foundation to help Australian students connect the dots from the basics to advanced topics in Genomics.
Apart from Genomics Assignment Help, our website also provides help with the various branches of biology.
The word Genomics is a confluence of the two terms: Gene – the hereditary unit and omics – the complete constituents.
Genomics deals with the study or analysis of all the genes of an organism. This study includes the structure, function, evolution, expression, interaction, mapping, and editing of genes.
Genomics is a multi-disciplinary approach. We understand genes through several branches of Biology.
These branches can be split into two types.
Based on how we study the genes, the branch of Genomics is categorised into three branches.
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Revolution in curing genetic diseases/abnormalities started with Gene Therapy in 1990. Gene Therapy supplements the functions of abnormal genes, through the therapeutic stem cells, with normal genes.
Gene editing technology started by the end of the 20th century, reached its full potential by 2015. Gene editing doesn’t supplement the functions of an abnormal gene but tries to rectify the abnormal gene. Rectification can be achieved through editing, modification, or deletion of the targeted gene.
Whatever the technology of Gene Editing, they all rely on inducing a Double Stranded Break (DSB) by programmed nucleases. DSBs are specific cuts at desired locations in the DNA. The cell has its own machinery to repair the induced DSB. Non-homologous end-joining (NHEJ) and Homologous-directed Repair (HDR) are the two ways in which DSB can be repaired.
NHEJ is imprecise and involves a range of proteins, that try to join the two broken ends of the DNA. This might involve some random insertions or deletions.
HDR is precise and uses a homologous copy of DNA (Donor Template) to correct the DSB.
Three frequently used tools that are used in gene editing technology include:
Gene Editing Technology
Zinc Finger Nucleases
Transcription Activator Like Effector Nucleases
Clustered Regularly Interspaced Short Palindromic Repeats
Zinc Finger Nucleases (ZFN):
Zinc Finger Nucleases are programmable restriction endonucleases. The two important components of ZFN are DNA binding Zinc finger domains and DNA-cutting domains.
DNA binding ZFNs can be modified artificially (programmed) to bind to target sites of the DNA. DNA cutting domains make cuts at the target sites.
The cuts are repaired by the cellular mechanisms, discussed above. For us to make a precise repair, along with ZFNs we add a donor DNA to generated DSB.
Transcription Activator Like Effector Nucleases (TALENs):
The TALEN technology uses two protein domains. TALE and FokI. TALE is DNA specific protein binding domain that resembles a transcription factor in function and structure, thus is the name Transcription Activator Like Effector (TALE). TALE is the programmable portion of TALEN.
FokI is an endonuclease, that requires prior binding of TALE to the DNA. FokI induces DSB.
As said before, the DSB’s are repaired. HDR requires a Donor DNA template.
CRISPR-CAS9 system was reported as an immune system in bacteria, against bacteriophages in 2007 and later transformed into a gene editing technology in 2015.
CRISPR (Clustered Regularly spaced short palindromic repeats) is a family of DNA sequences in lower microbes, the Bacteria and Achaea. These heritable sequences confer adaptive immunity to the bacteria and degrade the incoming viral RNA and DNA in bacteria.
When a phage infects a bacterium, the viral DNA is processed into pieces called protospacers (pS). The pS are inserted into the CRISPR locus. CRISPR locus has many direct palindromic repeats (shown in pink). pS are termed as Spacers (S), shown in green, once are inserted into the CRISPR locus. Each S is bound by palindromic repeats. CRISPR locus, therefore, serves as a record for the entry of many types of viruses that infect the bacteria.
When the same phage infects the bacteria the other time, the CRISPR-CAS system is activated to interfere with the viral replication. There is a region in bacterial DNA known as CRISPR-Cas Locus. This locus codes for a Cas system.
The Cas system has two components. A Cas protein and Guide RNA (gRNA).
The gRNA can come in two forms. The brown representation in the above figure is the Cas protein.
Forms of gRNA
Two Component gRNA (Red and Blue)
A crRNA (CRISPR RNA) and
One Component gRNA (Green)
crRNA and tracrRNA linked by linker loop
Cas Protein: Nuclease
crRNA: RNA of spacer and tracrRNA: Structural RNA, enhances the precision of Cas Protein.
The crRNA is complementary to the protospacer, whose adjacent motif is PAM (Protospacer Adjacent Motif). The Cas system binds to the target DNA and cleaves at the PAM site, creating DSB.
CRISPR is manipulated by programming the crRNA because crRNA has the ability to recognise the target sites.
The next thing that follows is the repair of DSB, which can happen in either of the two ways mentioned earlier.
1. Polymerase Chain Reaction (PCR) steps:
PCR is a technology that helps amplify the gene of interest / desired gene from nanomoles to millimoles.
PCR was developed in 1980 by Kary Mullis. PCR relies on an enzyme termed DNA polymerase. DNA polymerase uses a DNA as template and makes a billion copies of it. Taq polymerase is generally used to synthesise new DNA templates because it can operate at high temperatures.
The important ingredients in PCR include:
2 Primers (5’ – 3’) for two strands.
dNTPs (dATP’s, dTTP’s, dCTP’s and dGTP’s)
Taq Polymerase (Thermo aquaticus (taq) – the source of this enzyme).
PCR is a cycle and happens in three steps.
After Step 3, the newly formed DNA are cycled back to Step 1. PCR reactions are driven in an automated machine called as Thermocycler, that provides, appropriate temperatures for the individual steps. This cycling PCR can produce billions of DNA copies from hundreds of DNA.
2. Genome and Types of Genome:
The complete set of DNA of an organism is known as Genome.
Genomes can be categorised into two.
Eukaryotic Genomes can be further categorised into three.
Nuclear Genome: The complete set of chromosomes seen in a nucleus.
Chloroplast Genome: Chloroplasts are semi-autonomous organelles, majorly found in plants. The semi-autonomist character is attributed to the plasmid of the chloroplast. This plasmid is the Chloroplast genome.
Mitochondria Genome: Mitochondria are also semi-autonomous like Chloroplasts. The plasmid of Mitochondria forms the Mitochondrial Genome.
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