Questions tagged [cas9]

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4
votes
1answer
758 views

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 ...
4
votes
1answer
177 views

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 ...
3
votes
1answer
388 views

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 ...
2
votes
1answer
87 views

What is meant by “heads” and “tails” in the context of gene orientation?

I have a hard time understanding what this paper is talking about when it says: We observed maximal cleavage at sites oriented tail-to-tail and separated by -10 bp to +30 bp (Fig. 2d). Finally, ...
2
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0answers
849 views

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 ...
2
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0answers
74 views

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 ...
1
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0answers
66 views

Can CRISPR-cas9 be used to insert a large gene?

I would like to insert a 1500 bp or longer gene to the broken site in E.coli (or may be bacillus subtilis which support NHEJ) after cut with Crispr-Cas9. Is this possible for both NHEJ or HDR?
1
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0answers
118 views

Are exosomes useful as a transfection or delivery mechanism in gene editing?

The use of viruses as transfection or delivery agents for gene editing (CRISPR/cas9, etc) is well known. However, one problem with using viruses to deliver DNA into cells is the possibility of ...
0
votes
1answer
33 views

Can a protein go outside of nucleus after go inside with nuclear localization signal (NLS)

To make a protein (for example Cas9, GFP...) able to enter the nucleus, we need to add a NLS tag for it. So will it able to go outside the nucleus after get inside ?
0
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0answers
78 views

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 ...