Questions tagged [crispr]

Cluster Regularly Interspaced Short Palindromic Repeats; Naturally a prokaryotic adaptive immune system, elements of CRISPR are increasingly being used for sequence specific DNA targeting in biotechnological applications, especially genome editing. This tag can be used for questions about its natural function or biotechnological applications, including elements of the CRISPR system such as Cas9 and Cpf1 nucleases.

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Functions of tracrRNA and crRNA in the CRISPR/Cas9 system

I need some help about Crispr / Cas9. The CRISPR/Cas9 technic consists (for bacterias) in "cut" bacteriophage's DNA, in order to make it unfunctional. When the RNA is transcribed (RNA which is ...
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CRISPR/Cas9: What are the main differences between sgRNA and the Cr:TracrRNA ?

So from what I understand, in gene editing, the CRISPR vector expresses a small RNA sequence comprised of a small guide-RNA that is complementary to your target sequence. The sgRNA comprises a 20 Bp ...
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Why doesn't CAS9 cleave the original CRISPR sequence in the bacterial genome?

CAS9 is the RNA-guided endonuclease that cleaves DNA as specified by the RNA sequence and is used to target viruses that infect bacteria. So why doesn't this RNA+endonuclease combo also cleave the ...
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What is the role of tracrRNA in CRISPR-cas9?

From what I understand, in a CRISPR cas9 complex, gRNA is comprised of tracrRNA and crRNA. I've read that crRNA is the part which is matched to the DNA which is targeted, but what role does tracrRNA ...
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Is Cas9 unique in it's ability to act in response to a specific DNA sequence?

As a computer scientist, I'm interested in the ability of the Cas9 protein to function as an if-gate for DNA. It opens up so many questions for me. Right now the protein works as "if the sequence ...
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What´s the role or function of the homologous arms in a donor template in a knockout/knock edition via Crispr-cas9?

I have to make an exposition in my university about Crispr-cas9 edition and I have some questions about the method. In the knock out/knock in technique is used a plasmid containing the DNA that ...
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2answers
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How to edit/insert new gene after cutting with CRISPR/cas9

I'm a student started who has started learning about CRISPR/Cas9. As I understand it, CRISPR/Cas9 is an enzyme that is used to cut a gene at a specific sequence. I would like to know how scientist do ...
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Do users of CRISPR/Cas iterate or parallelize to try multiple guide sequences?

I've read about on-target efficiency and off-target effects in use of CRISPR/Cas9, and about tools that suggest good guide sequences. I am wondering: how many guide sequences do typical CRISPR users ...
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1answer
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How specific are CRISPR-cas9 cuts?

CRISPR-cas9 uses a string of RNA that matches with DNA and makes a double stranded cut at that point. If the RNA is just a few letters in length, the enzyme would cut DNA in many places. It would be ...
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why selecting cells after gene editing with CRISPR Cas9?

I am new to the CRISPR Cas9 genome editing system and I have the above basic question. Another way to think about this: what would happen if after cells are transfected, the cultures are left to grow ...
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1answer
200 views

What is the most up-to-date CRISPR/Cas9 protocol?

I am collecting literature to start a new project on CRISPR/Cas9 gene editing. I must put together a protocol to start and am intending to use the following paper as guidance: "Genome engineering ...
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Insertion of synthetic DNA sequence

If I have some synthetic DNA sequence (<=20 bp long), is there a way for me to reliably insert this sequence next to some n-bp motif? I'd like for this to be possible in humans. If so, are there ...
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In CRISPR bacteria, how does viral genomes get integrated into the spacers of CRISPR? Also, in its use, where does Cas9 cut the DNA?

I've been out of Biology for about a year polishing my programming skills. I know CRISPR/Cas9 allows targeted 'cutting' of DNA via RNA-guidance. Few questions regarding this. Regarding to its natural ...
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1answer
922 views

How does one insert cas9 into animal cells?

How could cas9 be inserted into cells by researchers looking to edit a genome? I imagine for engineering bacterial systems you could just put in the cas9 coding region in an expression vector, but is ...
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Why is dsDNA nuclease activity by CRISPR/Cas9 only shown indirectly?

In Jinek et al., the authors show nuclease activity of their CRISPR/Cas9 system using the so-called Surveyor assay method. This assay recognizes small mismatches in dsDNA which are introduced by error ...
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CRISPR-Cas Systems

In the context of the bacterial systems (not the gene editing tool), I was wondering what happens to the foreign DNA after the Cas proteins have created a new spacer. It is really not clear to me, ...
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239 views

Mutations/deletions with CRISPR

I need to stop some protein from being active and searching for some universal way to do so. In mammalians. With CRISPR it is possible to knock-out the entire gene. But it's a little complicate (...
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478 views

Splice in with CRISPR/Cas

I need to splice a gene into a human cell genome, with highest rate possible. I mean, doesn't really matter where the gene enters, nor does it matter if some cells die as a result of this. CRISPR ...
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0answers
102 views

CRISPR Knock in

Using the CRISPR/Cas9 technology, it is possible that after inducing a DSB with the Cas9 endonuclease guided with an RNA designed by the user and using a template DNA, get a desired Knock-In (KI) by ...

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