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How to deisgn sgRNA for your CRISPR?

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MaMaMia

Lvl 5
3y ago
Updated: 4/27/2022

CRISPR/Cas9 system consists of a Cas9 protein and a single guided RNA (sgRNA). The sgRNA could be divided into two parts: a 20 nucleotides targeting sequence and a scaffold sequence. The 20-nt sequence completely pairs to the genomic DNA and the scaffold RNA which is essential for Cas9 recognizing with sgRNA helps Cas9 bind to the genome. For this reason, Cas9 could recognize the target sequence and mediate a Double-Stranded Breaks (DSB) that located nearly 3 base pairs upstream of the Protospacer Adjacent Motif (PAM) sequence. Actually, PAM acts as the specific requirement for CRISPR/Cas9 systems that varies from different Cas9 orthologs, for example, PAM of S.Pyogenes Cas9 is 5’-NGG-3’ and Neisseria meningiditis Cas9 is 5’-NNNNGATT-3’. After DSBs are caused, Gene edition, no matter disruption or replacement, will be finished by gene repair mechanisms.

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MaMaMia

Lvl 5
3y ago

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Related Questions

What are the differences between crrna and sgrna in the context of genome editing techniques?

CRISPR RNA (crRNA) and single-guide RNA (sgRNA) are both used in genome editing techniques like CRISPR-Cas9. The main difference is that crRNA is a part of the natural CRISPR system in bacteria, while sgRNA is a synthetic molecule designed to combine the functions of both crRNA and tracrRNA. Both molecules guide the Cas9 enzyme to the target DNA sequence for editing, but sgRNA is more commonly used in research and applications due to its simplicity and efficiency.


What are the differences between sgRNA and crRNA in CRISPR technology and how do they impact gene editing efficiency?

sgRNA (single guide RNA) is a synthetic RNA molecule that combines the functions of both the crRNA (CRISPR RNA) and tracrRNA (trans-activating CRISPR RNA) in CRISPR technology. sgRNA simplifies the gene editing process by serving as a single molecule that guides the Cas9 enzyme to the target DNA sequence for editing. On the other hand, crRNA is a natural RNA molecule that specifically recognizes the target DNA sequence for editing. The use of sgRNA can improve gene editing efficiency by streamlining the process and reducing the risk of errors compared to using separate crRNA and tracrRNA molecules.


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