2022
DOI: 10.1186/s12864-022-08971-1
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A robust pipeline for efficient knock-in of point mutations and epitope tags in zebrafish using fluorescent PCR based screening

Abstract: Background Genome editing using CRISPR/Cas9 has become a powerful tool in zebrafish to generate targeted gene knockouts models. However, its use for targeted knock-in remains challenging due to inefficient homology directed repair (HDR) pathway in zebrafish, highlighting the need for efficient and cost-effective screening methods.  Results Here, we present our fluorescent PCR and capillary electrophoresis based screening approach for knock-in using… Show more

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Cited by 5 publications
(4 citation statements)
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“…This would not allow the changes we wanted to introduce into the alb or kcnj13 genes, T to G and T to A, respectively. Therefore, HDR is still a very convenient method for precise base changes in the zebrafish genome, especially when synthetic oligonucleotides can be co-injected as donor templates (Carrington et al, 2022). It is, however, desirable to find a target site for the endonuclease as close as possible to the position that should be changed.…”
Section: Discussionmentioning
confidence: 99%
“…This would not allow the changes we wanted to introduce into the alb or kcnj13 genes, T to G and T to A, respectively. Therefore, HDR is still a very convenient method for precise base changes in the zebrafish genome, especially when synthetic oligonucleotides can be co-injected as donor templates (Carrington et al, 2022). It is, however, desirable to find a target site for the endonuclease as close as possible to the position that should be changed.…”
Section: Discussionmentioning
confidence: 99%
“…Therefore, HDR is still a very convenient method for precise base changes in the zebrafish genome, especially when synthetic oligonucleotides can be co.injected as donor templates (Carrington, Ramanagoudr-Bhojappa et al 2022). It is, however, desirable to find a target site for the endonuclease as close as possible to the position that should be changed.…”
Section: Discussionmentioning
confidence: 99%
“…In the future it will be important to generate endogenously tagged Nrxn3 animal models to visualize whether Nrxn3 is present at ribbons synapses in hair cells. In recent years, adding endogenous tags to proteins in mice and zebrafish has become more straightforward, making this approach more straightforward (Carrington et al, 2022;Morrow et al, 2021). .…”
Section: Nrxn3 Interactions With Core Presynaptic Componentsmentioning
confidence: 99%