CRISPR based SHERLOCK Technology- The Detective!
For long, bacteria are using the natural biological system to protect themselves. One such major system that has been discovered lately by scientists is CRISPR. It is an enzyme which targets the foreign genetic material, cleaves it off and prevents viral or bacterial invasion. Scientists are modifying this natural system to fight many diseases and find out their cure. In light of this, researchers at Broad Institute have developed CRISPR based SHERLOCK technology to detect nucleic acids.
It is a breakthrough discovery. You must be wondering why? Well, though CRISPR based technology is well known for gene editing, that’s not the limit. The SHERLOCK technology can be used for diagnosis purposes of diseases like Zika, Ebola, and many more. The technology is highly sensitive and has proved to be very effective in disease diagnosis.
Mimicking the unique defense mechanism found in bacteria
Bacteria contain the genetic information of infectious agents in their genome at a particular location defined as CRISPR. When bacteria identify the presence of foreign invader either viral, bacterial, or any other infectious agent, it activates its immune system. The bacteria initiate the synthesis of unique RNA, also known as guide RNA. This guide RNA complexes with Cas9 enzyme (CRISPR associated protein 9), which is an endonuclease enzyme that cuts the DNA at certain target sites. This mechanism of defense is used for the development of SHERLOCK technology.
What is SHERLOCK technology?
The Cas9 enzyme is also known as a programmable nuclease. This is due to the fact that one can use it to cut the targeted DNA site by providing a unique RNA. This method is harnessed further by scientist to develop SHERLOCK technology (stands for Specific High sensitive Enzymatic Reporter Unlocking). The only difference is that it uses Cas13a endonuclease enzyme rather than Cas9. The method involves the following:
- RNA amplification (Or DNA by enzyme reverse transcriptase).
- Amplified RNA is complexed with Cas13a enzyme, a guide RNA resembling target sequence, and a reporter nucleic acid sequence containing a fluorescent tag.
- Activation of Cas13a non-specific RNAse activity in response to the presence of the target sequence is detected in the reaction mixture
- As a result, the RNA reporter will be cleaved leading to activation of the fluorophore.
Since the technique is based on fluorescence-based detection of the target sequence from the pool of amplified nucleotides, it is known as “Specific High sensitive Enzymatic Reporter Unlocking” i.e. SHERLOCK.
Are there other similar technologies?
- DETECTR Technology (DNA Endonuclease Targeted CRISPR Trans Reporter):
This system was first reported by scientists – Jennifer Doudna and her colleagues at the University of California. This technology is similar to SHERLOCK with a difference in the type of endonuclease used. Here the researchers used Cas12a instead of Cas13a or Cas9 which cleaves the dsDNA and leads to activation of the non-specific ssDNA breakdown. The researchers used this technique to identify carcinomas related Human Papilloma Virus (HPV).
- HUDSON Technology (Heating Unextracted Diagnostic Samples to Obliterate Nuclease):
This method involves the release of nucleic acids directly into the solution by initiating the heat and chemical reduction based lyses of the viral envelope. The researchers have combined the HUDSON technology with SHERLOCK technology in order to detect two most fatal virus i.e. Zika and Dengue virus.
How SHERLOCK Technology can be helpful?
As we have looked upon the working mechanism of the technology, it is easy to guess its enormous potential in the field of biosciences. To mention the few:
- It can be used for detection of both viral and bacterial pathogens
- The evaluating viral load of HIV patients
- Detection of any DNA mutation and subsequent assessment of disease tendencies
- Rapid and precise result for TB (Tuberculosis)
- Instantaneous identification of pathogen-resistant genes
- Assessment of gene edits created by CRISPR using SHERLOCK technology
CRISPR based technologies are hitting the science news headlines almost every day. This shows the exponential prospects the technology has in the future. The same applies to SHERLOCK technology being a highly sensitive, specific, simple and flexible technique it has many applications. There is no doubt that soon there will be many more such developments that can help mankind to overcome the biological complexities of the future. Till then let’s ponder upon the existing technology, analyze them, and evaluate their competencies to come up with something new.