2. CRISPR
CRISPRs (clustered regularly interspaced short palindromic repeats) are repetitive nucleotide sequences followed by a short spacer DNA segments integrated into the genomes of many bacteria and archaea upon exposure to invading pathogens.

3. CRISPR Clustered Regularly Interspaced Short Palindromic Repeats
originally discovered in the bacterium Streptococcus pyogenes
mechanism to defend against viruses and foreign DNA
has provided a revolutionary tool for targeted genome engineering with simple elegance

6. Cas9
Cas9 (or "CRISPR-associated“ protein) is an enzyme that acts like a pair of molecular scissors, capable of cutting strands of DNA.
Ribonucleoprotein (RNP)

7. CRISPR-associated (Cas) nucleases form the CRISPR-Cas machinery to rid the prokaryotes of foreign genomic elements (target DNA).

8. Cas9 Nuclease

9. Once the Cas9 protein is activated, it stochastically searches for target DNA by binding with sequences that match its protospacer adjacent motif (PAM) sequence. A PAM is a two- or three-base sequence located within one nucleotide downstream of the region complementary to the guide RNA.
When the Cas9 protein finds a potential target sequence with the appropriate PAM, the protein will melt the bases immediately upstream of the PAM and pair them with the complementary region on the guide RNA. If the complementary region and the target region pair properly, the Cas9 will cut the target DNA after the third nucleotide base upstream of the PAM

11. Cas Genome Editing

13. How CRSPR Works

14. In 2012, CRISPR was utilized in eukaryotes and a readily-programmable two-component CRISPR system was used to disrupt targeted genomic sites.

15. The CRISPR-Cas9 system can make genetic manipulations much faster than previous used methods. Instead of years to make a genetically engineered mouse, it now can take as little as 4 months!

16. Applications of Crispr
Eradication of genetic disease
Eradication of infectious disease
Eradication of invasive species
Increase life expectancy
Designer Babies
Super Soldiers

18. CRISPR-Cas-system-Adaptive-immunity-

19. The invading DNA is fragmented and a new protospacer (green) is selected, processed and integrated as a new spacer at the leader end of the CRISPR array. During the second stage, which is known as expression, the CRISPR locus is transcribed and the pre-crRNA is processed into small crRNAs by CRISPR-associated (Cas6) and/or housekeeping ribonucleases (such as RNase III). The mature crRNAs and Cas proteins assemble to form a crRNP complex. During the final stage of interference, the crRNP scans invading DNA for a complementary nucleic acid target and on successful recognition, the target is eventually degraded by Cas nucleases.

21. The recognition and degradation of invasive DNA by CRISPR/Cas type II occurs in three steps:
Acquisition: Invasive DNA is recognized via a protospacer adjacent motif (PAM) – the sequence NGG. A short sequence downstream of the PAM sequence is then integrated into the host CRISPR array and is termed spacer. Spacer sequences transcribe for CRISPR RNAs(crRNAs) which help to cleave sequence-specific invasive DNA. These sequences are located between short palindromic repeats, which are neccessary for the functionality of the crRNAs.
Expression/Transcription: The Cas9 endonuclease is expressed. CRISPR array is then transcribed and processed by RNAse III into crRNAs. These contain the complementary spacer sequence and the direct repeat sequence. The crRNA guides the Cas9 protein specifically to invasive DNA sequences. Furthermore trans-activating crRNAs (tracrRNA) are transcribed and bind to the direct repeat part of the crRNA. The tracrRNA is necessary for the formation of a Cas9-RNA complex.
Interference: Repeatedly invading DNA, which has been integrated into the CRISPR locus, is detected by the RNA-protein complex and cleaved by Cas9.

22. CRSPR and Gene drives against Malaria