CRISPR Array

Once the complex was bound to the DNA, a cut would be made to eliminate and destroy the invaders. 83% of archaeal genomes and 45% of bacterial genomes (Shabbir, M. et al, 2016) were shown to be able to successfully utilize the CRISPR Cas9 system. These are very promising statistics, so it is no wonder that there has been such an advancement in the past few years to bring this technology to eukaryotic cells, mammalian cells and eventually human cells.

CRISPR stands for Clustered Regularly Interspaced Short Palindromic Repeat, referring to the repeating DNA sequences found in the genomes of microorganisms. CRISPR technology allows scientists to make precise changes in genes by splicing and replacing these DNA sequences with new ones. Through these changes, the biology of the cell is altered and possibly affects the health of an organism. The possibilities are endless as this offers opportunities in curing deadly diseases, modifying genes, and changing humanity as we know it. Although bioengineering has been around since the 1960s, CRISPR is significant because of the comparative low costs and the ease of the procedure to

The CRISPR Team was fortunate to be a part of the “Virus Documentary” (SciChannel) and conduct successful experiments discovering the activity of viruses. Through a series of test conducted by The CRISPR Team, it

The antibiotic-resistant population that are not re-sensitised can consequently be displaced by lytic phages. Yosef and colleagues cultured bacteria containing ndm-1 and ctx-M-15 plasmids conferring resistance to carbapenems, a -lactam antibiotic. These cultures were treated with a lysogenizing phage cas-CRISPR, or a control cas phage, followed by the addition of T7-N1C1 lytic phage of which the lysogenized cas-CRISPR bacteria were protected. A significantly greater number of the bacterial cells treated with cas-CRISPR phage were resistant to the T7-N1C1 phage as compared to those cells treated with the control cas phage (Yosef, et al., 2015). To confirm the loss of antibiotic-resistant genes, phage-resistant colonies were inoculated onto streptomycin-containing plates, revealing that all bacteria lysogenized with cas-CRISPR phage lost their resistance and became sensitive to streptomycin (Yosef, et al., 2015). Therefore, Yosef successfully demonstrated that selective pressure for antibiotic sensitive bacteria can be achieved through the use of phages. However, sensitised bacteria could be less fit than resistant bacteria due to the genetic burden of the transferred CRISPR-Cas and accompanying phage genes.

CRISPR or Clustered Regularly Interspaced Short Palindromic Repeat, is used to change the DNA. Today, as humans, we have learned how to use CRISPR for what we want it to do. This is a major break in what we know about DNA. For the future we are looking at how we can change DNA and control what the DNA changes to.

With the use of CRISPR, a specific gene in the genome of a cell can be targeted and mutated to rid of the preexisting mutation. The technique works through the use of an enzyme called Cas9, which acts as the “scissors” to cut two strands of DNA at a specific location in the genome to allow for pieces of DNA to be added or removed. Another molecule of use in the process is a piece of RNA called guide RNA (gRNA). Guide RNA comprises of a small piece of pre-designed RNA sequence located within a longer RNA strand. This longer RNA strand binds to DNA and the pre-designed sequence guides the Cas9 to the correct part of the genome. This occurs for the assurance that the Cas9 enzyme cuts the right part of the genome out. The article provides specific cancers and genetic diseases and the targets for CRISPR/Cas9 that act on these mutations. Amongst the cancers, lung, thyroid, and breast cancer was mentioned. The genetic diseases mentioned were Huntington disease, Alzheimer’s and muscular

Ryland F. Young III’s article, “Secret Weapon,” begins by explaining the efficiency of CRISPR defense. Bacteria’s are infected by bacteriophages 1030 times a day. Bacteriophages mutate and undergo an unfathomable amount of genomic recombination that makes it difficult for bacteria to defend themselves from these viruses. Scientist discovered a new bacteria defense systems called CRISPR. This process in combination restriction-modification, restriction enzymes and apoptosis allows bacteria to protect themselves from invading viruses. CRISPR defense is effective because it forced bacteriophages to undergo intense recombination shuffling down to a scale of the size of CRISPR spaces, instead of just mutating. Though CRISPR defense is an effective

infection by viruses. When a bacterium identifies a threat from a virus, it generates two types of

Even though CRISPR/Cas9 was just discovered around 4 years ago, the CRISPR/Cas system itself was first observed in bacteria (E.coli). It was what CRISPR was doing in the bacteria that makes it so unique. CRISPR stands for, “clustered regularly interspaced short palindromic repeats”. This means that there are going to be short sequences of DNA bases that will read the same forward and backwards. In-between these short sequences will be spacer DNA. Unlike the palindromic sequences, the spacer DNA will have sequences

Scientist Francisco Mojica was the first scientist to characterize a CRISPR locus in 1993, at the University of Alicante in Spain. Mojica studied CRISPR throughout the 1990’s and early 2000’s, and in 2002 the term CRISPR was first used in print by Ruud Jansen. Later, in 2005, Mojica discovered that sequences he discovered matched sections of genes from bacteriophage, which led him to hypothesize that CRISPR is actually an immune system from the bacteriophage. His hypothesis was indeed correct. Also in 2005, Cas9 was discovered by Alexander Bolotin, at the French National Institute for Agricultural Research while he was researching recently sequenced bacteria. Bolotin found that a CRISPR locus was similar to others that had already been discovered,

This easy-to-use technique will facilitate understanding genome functions and their relationships. It has sparked a revolution in genome engineering field since 2012. Below we review the history of CRISPR-Cas9 system development, reveal its underlying molecular mechanism and discuss its applications, challenges and future avenues of this novel

Thus, spacers operate as a ‘genetic memory’ of the preceding infections. If an infection caused by the same virus were to occur, the defense system of CRISPR would cut any DNA sequence that matches the spacer sequence, and will consequently protect the bacterium from any viral attacks. If an earlier undetected virus attacks, a new spacer will be created and added to the repeats and spacer chain. The three basic steps that the CRISPR immune system follows to protect bacteria from repeated viral attacks are as follows. Step 1 is adaptation, where the DNA from the attacking virus is managed into short segments, which are injected as new spacers into the CRISPR sequence. Step two is the production of CRISPR RAN, where the CRISPR spacers and repeats that are in the bacterial DNA go through transcription, the method of copying a segment of DNA into RNA. The resultant is a single-chain RNA molecule, which is cut into shorter segments that are called CRISPR RNAs. Step three is targeting, where the CRISPR RNAs will direct the bacterial molecular machinery to terminate the viral

The question that should be asked now is how the CRSIPR sequences simply defend the micro-organism from invading viruses. In figure one; below there is a sequence of black diamonds (which represent DNA) and in between them are spacers. This is what is known as a CRISPR sequence. The spacers are vital for the immunity of the micro-organism or bacteria. They contain the genetic information of the previous viruses that have attacked this micro-organism. Furthermore, when a new virus attacks the micro-organism, a new spacer is added to the CRISPR sequence. This allows the immune system to recognise and silence exogenous genetic material. The process of this happens as the DNA of the virus is cut into short segments by the Cas (CRISPR associated)

The RNA-guided CRISPR-Cas9 system has revolutionized genetic engineering by allowing targeted genomic modification through the simple design of a 20 base pair guiding sequences. This groundbreaking technology utilizes a short guide RNA (sgRNA) to direct the Cas9 nuclease to a specific genomic locus through complementary base pairing. The guide RNA or sgRNA is responsible for the specificity of the CRISPR-Cas9 system, and several consideration need to be taken during its design process. In bacteria and archaea, the crRNA and tracrRNA form a complex which acts as the homing device for directing the Cas9 nuclease to the invading foreign genetic materials. The tarcrRNA 's scaffolding ability along with the specificity of the crRNA can be

What is CRISPR? As mentioned above it is the clustered regularly interspaced short palindromic repeats, which are segments of DNA that have short repetitions of base sequences. Each repetition is followed by short segment, which is referred to as spacer DNA. CRISPR is actually found in bacteria and archaea and occurs naturally. Turns out CRISPR is part of the immune systems for bacteria, in those spacer DNAs are store genetic information of viruses that have attacked the bacteria. CRISPR has the ability to capture and store genetic information from invading viruses and use it as template in order to fight off the virus. Along with CRISPR, they have enzymes called CRISPR-associated (Cas) that aid CRISPR in destroying any recognizable virus. The mechanism for CRISPR/Cas goes through this