Remerciez-le!

Remerciez @Admin pour avoir partagé cet document gratuitement, de la manière la plus simple, en partageant sur les réseaux sociaux.

University of Management and Technology _______________________________________

University of Management and Technology __________________________________________________ ____________________________________________________________________ Name: Seerat Rani ID: F2019231060 Section: A2 Submitted To: Sir Muhammad Khurram Topic: CRISPER/Cas9:Gene Editing Write a summary? CRISPER/Cas9: Gene Editing: CRISPR/Cas9 a genome editing technology derived from a bacterial innate immune system found in variety of bacteria and archaea, are fragments of prokaryotic DNA containing short repetitions of base sequences. (Jiang& Doudna, 2017)CRISPR/Cas9 involves two essential components: a guide RNA called synthetic single guide RNA (sgRNA) is the section of RNA which binds to the genomic DNA of 18–20 nucleotides to match a desired target gene, and Cas9 (CRISPR-associated protein 9)(Redman et al., 2016)an RNA-guided endonuclease from microbial adaptive immune system clustered regularly interspaced short palindromic repeats (CRISPR) which causes a double-stranded DNA break, allowing modifications to the genome. It makes it possible to correct errors in the genome and turn on or off genes in cells and organisms quickly, cheaply and with relative ease.(Yusof, 2018) The CRISPR array is made up of an AT-rich leader sequence followed by short repeats that are separated by unique spacers. CRISPR repeats typically range in size from 28 to 37 base pairs (bps). Some show dyad symmetry, implying the formation of a secondary structure such as a stem-loop ('hairpin') in the RNA. CRISPR-Cas systems have two classes. Class 1 systems use a complex of multiple Cas proteins to degrade foreign nucleic acids. Class 2 systems use a single large CAS protein for the same purpose. Cas9 features a bi-lobed architecture with the guide RNA nestled between the alpha-helical lobe and the nuclease lobe. These two lobes are connected through a single bridge helix. (Bannikov& Lavrov, 2017)There are two nuclease domains located in the multi-domain nuclease lobe, the RuvC which cleaves the non-target DNA strand, and the HNH nuclease domain that cleaves the target strand of DNA. The RuvC domain is encoded by sequentially disparate sites that interact in the tertiary structure to form the RuvC cleavage domain.A key feature of the target DNA is that it must contain a protospacer adjacent motif (PAM) consisting of the three-nucleotide sequence- NGG. (Jiang& Doudna, 2017) Adaptive immunity occurs in three stages: (i) insertion of a short sequence of the invading DNA into the CRISPR array as a spacer sequence (ii) transcription of precursor CRISPR RNA (pre-crRNA) which goes through maturation to yield individual crRNAs, each composed of a repeat portion and an invader-targeting spacer portion (iii) crRNA-directed cleavage of foreign nucleic acid because of Cas proteins at sites complementary to crRNA spacer sequence Three different types of CRISPR/Cas system had been identified; Type I, II and III that utilizes different mechanism in generating CRISPR RNA (crRNA) and CAS proteins. For the purpose of genome editing, researchers (Redman et al., 2016) adapted the Type II system based on its simplicity in requiring only CRISPR-associated protein 9 (Cas9) proteins from Streptococcus progenes and two other RNA components, crRNA and trans-activating crRNA (tracrRNA) that fused to become guide RNA (gRNA). (Hille &Charpentier, 2016)In this system, the tracrRNA which is complementary to the repeat sequences in pre-crRNA triggers processing by double- stranded RNA-specific ribonuclease, RNase III in the presence of Cas9 protein. The gRNA then binds to Cas9 protein and direct the newly formed complex to a target sequence in the genome based on complementary base-pairing rule that allows a stretch of ~18 to 20 nucleotides of the gRNA sequence to hybridize with the targeted sequence, docking the Cas9 nuclease at that location. (Meiliana et al., 2017) Cas9 that consist of two different domains; HNH and RuvC-like, requires a base-pairing structure forming between the activating tracrRNA and targeting crRNA (gRNA) to cleave the double stranded DNA (dsDNA). Locus of site-specific cleavage in the genome is determined by base-pair mediated binding to complementary DNA sequences between crRNA (of the gRNA structure) and the target sequence. The target sequence must be adjacent to a protospacer adjacent motif (PAM), a short motif consisting of ‘NGG’ or ‘NAG’ sequence. The dsDNA strand complementary to the target-binding crRNA sequence is cleaved by Cas9 HNH domain while the non-complementary DNA strand is cleaved by Cas9 RuvC-like domain at a site three base pairs upstream of the PAM.(Jiang& Doudna, 2017)The DSB generated by Cas9 will induce the cell to repair the damage by either NHEJ or HDR. This is the focus point where researchers can manipulate the CRISPR/Cas9 system to modify the genome at targeted locations. One of the most exciting applications of CRISPR/Cas9 is its potential use to treat genetic disorders caused by single gene mutations. Examples of such diseases include cystic fibrosis (CF), Duchenne’smuscular dystrophy (DMD) and haemoglobinopathies. It’s used to treat infectious diseases, such as HIV. Although antiretroviral therapy provides an effective treatment for HIV, no cure currently exists due to permanent integration of the virus into the host genome. Showed as the CRISPR/Cas9 system could be used to target HIV-1 genome activity. (Christopher et al., 2018) There has been increasing interest in the possibility of using CRISPR/Cas9 to modify patient-derived T-cells and stem/progenitor cells which can then be reintroduced into patients to treat disease. There is also interest in using CRISPR/Cas9-mediated genome editing in pluripotent stem cells or primary somatic stem cells to treat disease. (Christopher et al., 2018) sTools from genome modifiers like clustered regulation to interspersed short pelindomic repeat (CRISSPR) - Proportional system (CAS) widely used to quote genes in animal zygotes and human cell model systems To do It's about the ultimate waste for basic and basic research and clinical applications. To date, DIC procedures have been consulted in early human embryos, and there is still some time left in the effective use and potential targeting of technologies such as CRISPR / CAS9. In this report, we use tripronuclear (3PN) zygotes in human cells, ranging from further investigations of CRISSPR / CAS9- mediated gene modification. Know-how to stop the globin gene (HBB). However, HBB's Homosexual Guide (HDR) is low performance and modified fetal mosaic. Off in T3E zygotes The outbreak of the target was also clear, as shown by the T7E1 payment and the complete configuration. In addition, the endogenous delta-globin gene (HBD), which is becoming compatible with HBB, has to contend with external donor oligos, which is a good thing. Our statistics also do not indicate that the genies in which they are objects of BB lux are in the place of non-cross and HD art. Together, our work highlights the seriousness of the CSSPR / CAS9 platform and the need to further improve the selection of coatings, CRSIPR / CAS9 arbitration modification for any medical application. Is a condition. Is. Our statistics also do not indicate that the genies in which they are objects of BB lux are in the place of non-cross and HD art. Together, our work highlights the seriousness of the CSSPR / CAS9 platform and the need to further improve the selection of coatings, CRSIPR / CAS9 arbitration modification for any medical application. Is a condition. Is. Our statistics also do not indicate that the genies in which they are objects of BB lux are in the place of non-cross and HD art. Together, our work highlights the seriousness of the CSSPR / CAS9 platform and the need to further improve the selection of coatings, CRSIPR / CAS9 arbitra He has significant potential in genome engineering and gene therapy, making the CRSPR / CAS9 system an important tool in biomedical research for many applications. Has emerged as To achieve conditional control of the CRISPR / CAS9 system, the genetically encoded light-activated CAS9 was engineered through a site-related installation of the Panjrad Licensed Amino Acid. Numerous license residues were identified as viable caging sites that could be optically modified to control the CAS9 function, such as by disabling both optical activation and external and endogenous gene function. Has been revealed. 1tion modification for any medical application. Is a condition. Death-dependent β-thalassemia (TDT) and scale cell disease (SCD) are acute monogenic diseases with severe and potentially life-threatening manifestations. BCL11A is a replicating factor that suppresses glo-globin expression and fetal hemoglobin in erythroid cells. We perform electrophoresis of CD34 + hematopoietic stem and progenitor cells obtained from healthy donors, including CRISPR-Cas9, targeting a specific BCR11A erythroid supplement. About 80% of the alleys in this locus were modified, with no evidence of invalid editing. After undergoing myeloabulation, two patients - one with TDT and the other with SCD - obtained modified autologous CD34 + cells by CTRPR-Case 9 containing the same BCL11A. The enhancer was targeted. After more than a year, bone marrow and blood clotting were higher in both patients There was an increase in the level of fetal hemoglobin that was distributed, freedom of transmission, and elimination of vaso-event episodes (in patients with SCD). (CRISPR is funded by Therapeutics and Vortex Pharmaceuticals for clinical trials. Gov numbers, NCT03655678 for CLIMB THAL-111 and NCT03745287 for CLIMB SCD- 121Because of its compatibility with gene editing research, predicting change from target to change is a hot topic in CRISPR-CAS9. Current forecasting methods have been developed. However, most of them only calculated uploads/s3/ seerat-rani-f2019231060-biochem-lab.pdf