Genome editing in plants with CRISPR/Cas9 and future perspective of CRISPR/CAS9

Genome editing with CRISPR/Cas9 technology can be used to make precise changes to the DNA of plants. This can be used to introduce new traits or characteristics into the plant, such as increased resistance to pests and diseases, improved crop yields, or enhanced nutritional content.

One of the benefits of using CRISPR/Cas9 for genome editing in plants is that it is a relatively simple and efficient process. Scientists can use the technology to make specific changes to the genome of a plant in just a few steps.

First, the researchers must design a guide RNA that will bind to the target DNA sequence that they want to modify. They can then use this guide RNA, along with the Cas9 enzyme, to cut the DNA at the desired location. Once the DNA has been cut, the plant’s natural repair mechanisms will kick in, and the DNA will be repaired by either replacing the cut section with a new DNA sequence or by filling in the gap with the same DNA sequence as before.

There are many potential applications for genome editing in plants using CRISPR/Cas9 technology. For example, scientists could use it to develop crops that are more resistant to pests and diseases, which could help to reduce the need for pesticides and other chemicals in agriculture. Genome editing could also be used to improve the nutritional content of crops, making them more nutritious and beneficial for human health. However, there are also many ethical concerns surrounding the use of genome editing in plants, and more research is needed to understand the long-term impacts of these changes.

Who discovered CRISPR?

CRISPR (clustered regularly interspaced short palindromic repeats) was first discovered in the late 1970s by two independent groups of researchers. In 1987, Salvador Luria and his colleagues at the University of Milan published a paper describing the discovery of CRISPR in the bacteria Escherichia coli. Around the same time, another group of researchers led by Ying-Hsiu Su at the University of California, Berkeley also published a paper describing the discovery of CRISPR in E. coli.

However, it wasn’t until the early 2000s that researchers began to realize the potential of CRISPR as a tool for gene editing. In 2005, researchers at the University of California, Berkeley, and the University of Utah published a paper describing the use of CRISPR to knock out specific genes in bacteria. This marked the beginning of the CRISPR revolution, and since then, CRISPR has become a widely used tool for gene editing in many different fields.

In 2012, Jennifer Doudna and Emmanuelle Charpentier published a paper describing the use of CRISPR to edit genes in human cells, which marked a major milestone in the development of CRISPR as a gene editing tool. Since then, their work has been recognized with numerous awards and accolades, including the Nobel Prize in Chemistry in 2020.

How does CRISPR work step by step?

CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) is a powerful tool for editing genes, which allows researchers to make precise changes to the DNA of living organisms. Here is a step-by-step breakdown of how CRISPR works:

1.  First, scientists identify the specific gene or genes that they want to target for editing.
2.  Next, they design a small piece of RNA called a guide RNA (gRNA) that is complementary to the target gene sequence. The gRNA consists of a short stretch of RNA with a specific sequence that will bind to the target DNA sequence.
3. The gRNA is then combined with a nuclease enzyme called Cas9, which acts as a pair of “molecular scissors” that can cut DNA.
4. The gRNA-Cas9 complex is introduced into the cells of the organism being edited, either through traditional genetic engineering techniques or using a virus to deliver the complex into the cells.
5. Once inside the cells, the gRNA guides the Cas9 enzyme to the target gene. The gRNA-Cas9 complex then cuts the DNA at the specific location specified by the gRNA.
6. At this point, there are a few different things that can happen, depending on the specific goal of the researchers:

• If the goal is simply to disable the function of the target gene, the researchers can leave the DNA cut as is. The cell’s natural repair processes will then kick in and try to repair the cut, but they may not do so perfectly, resulting in a mutation that disables the gene.

• If the goal is to insert a new piece of DNA at the cut site, the researchers can provide the cell with a template for the desired new sequence. The cell’s repair processes will then use this template to insert the new sequence into the genome at the cut site.

• If the goal is to delete a piece of DNA, the researchers can simply wait for the cell’s repair processes to attempt to repair the cut. If the cell doesn’t have a matching sequence to use as a template, it will simply delete the piece of DNA between the two cuts.

• Once the editing is complete, the modified cells can be returned to the organism, or they can be cultured in a dish to create modified cell lines or tissues.

CRISPR has revolutionized the field of gene editing, making it faster, cheaper, and more precise than ever before. It is being used in a wide range of research and therapeutic applications, including the development of new drugs, the study of disease mechanisms, and the development of genetically modified crops.

Future perspective of CRISPR/CAS9 gene editing:

CRISPR gene editing has the potential to revolutionize the field of plant biology and agriculture. It can be used to improve crop yields, reduce the need for chemical pesticides and fertilizers, and develop new plant varieties that are resistant to diseases, pests, and environmental stressors.

One potential application of CRISPR in plants is to create crops that are more resistant to drought, heat, and other extreme weather conditions. This could help to increase food security in regions that are affected by climate change. CRISPR could also be used to develop crops that are more nutritious, such as rice with higher levels of vitamin A or corn with enhanced protein content.

Another potential use of CRISPR in plants is to create biofuels from crops that are more efficient at converting sunlight into energy. This could help to reduce our reliance on fossil fuels and mitigate the greenhouse gas emissions that contribute to climate change.

Overall, the future of CRISPR gene editing in plants looks bright, with the potential to significantly improve agriculture and food security around the world. However, it is important to carefully consider the risks and ethical implications of this technology as it is developed and used.

What problems can CRISPR solve?

Medicine: CRISPR can be used to edit the genes of human cells to treat genetic diseases such as sickle cell anemia and cystic fibrosis. It can also be used to create animal models of human diseases to help researchers understand and develop treatments for these conditions.

Agriculture: CRISPR can be used to modify the genes of crops to make them more resistant to pests, diseases, and environmental stresses, which can increase crop yields and improve food security. Environmental conservation: CRISPR can be used to edit the genes of invasive species to reduce their impact on native ecosystems, or to restore the genetic diversity of endangered species.

Industrial applications: CRISPR can be used to modify the genes of microorganisms to produce valuable products such as drugs, fuels, and chemicals. Overall, CRISPR has the potential to solve a wide range of problems in various fields by allowing scientists to make precise edits to the genome of living organisms.

Constraints:

It is important to be aware of its limitations and constraints.

Off-target effects: CRISPR can sometimes make unintended changes to the genome, known as off-target effects. This can occur when the CRISPR guide RNA (gRNA) targets a sequence that is similar to the intended target, but not identical. Off-target effects can be minimized by using high-quality gRNAs and by using computational tools to predict and avoid potential off-target sites.

Incomplete editing: CRISPR may not always make a complete edit to the genome, and may leave behind incomplete or partial edits. This can be a problem when the goal is to delete or replace a specific sequence.

Insertion or deletion (indel) biases: CRISPR is more likely to insert or delete (indel) base pairs when making edits to the genome. This can be a problem if the goal is to make a precise point mutation, as the indel bias may introduce additional changes to the sequence.

Editing efficiency: The efficiency of CRISPR editing can vary depending on the target site and the organism being edited. In some cases, CRISPR may not be able to make a change to the genome at all.

Ethical considerations: The use of CRISPR to edit the genome of humans, plants, and animals raises ethical concerns, including the potential for unintended consequences and the potential for the technology to be used for non-therapeutic purposes. These concerns are being actively debated by scientists, ethicists, and policymakers.

The technology has the potential to improve our understanding of genetic diseases, develop new treatments for a wide range of diseases, and improve crop yields in agriculture.

There may also be potential for CRISPR to be used in clinical settings in Pakistan, although this will depend on the development of regulatory frameworks and the availability of trained professionals to use the technology safely and effectively.

It is important to note that the use of CRISPR also raises ethical concerns, and it will be important for Pakistan to carefully consider these issues as the technology is developed and implemented.