Recent Advances in CRISPR/Cas9 Delivery Strategies

Yip, Bon Ham

doi:10.3390/biom10060839

Cited by 228 publications

(236 citation statements)

References 103 publications

(189 reference statements)

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“…In addition to the health sector, CRISPR–Cas9 has great potential to affect agriculture and is used to expedite livestock and crop breeding [ 138 ]. On the other hand, there are numerous controversies related to CRISPR, including off-target effects, the immunogenicity of Cas9 nucleases and carcinogenic effects of CRISPR components, which require exhaustive analysis and scientific explanations [ 139 ].…”

Section: Discussionmentioning

confidence: 99%

Various Aspects of a Gene Editing System—CRISPR–Cas9

Janik

Niemcewicz

Ceremuga

et al. 2020

IJMS

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The discovery of clustered, regularly interspaced short palindromic repeats (CRISPR) and their cooperation with CRISPR-associated (Cas) genes is one of the greatest advances of the century and has marked their application as a powerful genome engineering tool. The CRISPR–Cas system was discovered as a part of the adaptive immune system in bacteria and archaea to defend from plasmids and phages. CRISPR has been found to be an advanced alternative to zinc-finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN) for gene editing and regulation, as the CRISPR–Cas9 protein remains the same for various gene targets and just a short guide RNA sequence needs to be altered to redirect the site-specific cleavage. Due to its high efficiency and precision, the Cas9 protein derived from the type II CRISPR system has been found to have applications in many fields of science. Although CRISPR–Cas9 allows easy genome editing and has a number of benefits, we should not ignore the important ethical and biosafety issues. Moreover, any tool that has great potential and offers significant capabilities carries a level of risk of being used for non-legal purposes. In this review, we present a brief history and mechanism of the CRISPR–Cas9 system. We also describe on the applications of this technology in gene regulation and genome editing; the treatment of cancer and other diseases; and limitations and concerns of the use of CRISPR–Cas9.

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Section: Discussionmentioning

confidence: 99%

Various Aspects of a Gene Editing System—CRISPR–Cas9

Janik

Niemcewicz

Ceremuga

et al. 2020

IJMS

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“…Virus vectors have been successfully employed for editing the animal cell genome by delivering the CRISPR/Cas9 system both in vivo and in vitro [26]. Viruses have also been utilized in plants for virusinduced gene silencing (VIGS) by introducing small interfering RNA and have become a promising tool for DNA-free plant genome editing [27,28].…”

Section: Crispr/cas Delivered As Virus-like Particlesmentioning

confidence: 99%

“…Recent advances in CRISPR/Cas delivery strategies and approaches were summarized in several excellent reviews recommended for further reading [2,26,36,38,44,45]. It should be noted that these articles are not focused entirely on DNA-free delivery but on the genome editing as whole.…”

Section: Emerging Trendsmentioning

confidence: 99%

DNA-free gene editing in plants: a brief overview

Tsanova

Stefanova

Topalova

et al. 2020

Biotechnology & Biotechnological Equipment

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The conversion of bacterial CRISPR/Cas defense system into a simple and efficient tool for genome manipulations brought experimental biology into new dimensions. Suddenly, genome editing reached many groups most of which were interested in it but not able to employ the available time-and labor-consuming approaches of the pre-CRISPR era. In plant biology and biotechnology, CRISPR/Cas gene editing became the second most important technology after plant transformation. Actually, it relies on the available array of methods of gene delivery. While sufficient for most purposes, the classic gene transfer methods might become a problem for some experimental settings. The main obstacle is that they include DNA delivery and, frequently, its subsequent integration into cellular genome. For this reason novel methods to achieve gene editing without the need of stable transformation and even without DNA delivery were developed. These new approaches include in vitro ribonucleoprotein complexes formulations (delivered by microinjection, particle bombardment, electroporation, liposomes etc.), use of virus-like particles and employment of bacterial secretory systems for Cas/gRNA delivery. The first attempts to achieve DNA-free editing were made less than ten years ago. Later, different types of animal and plant cells were addressed. In this mini review we try to summarize the current developments and emerging trends in the field of DNA-free editing in plants.

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“…However, it has proven challenging to induce cells to utilize donor DNA to conduct homology-directed repair (HDR), resulting in non-homologous end joining (NHEJ) repair, which is error-prone 3 . To date, the most efficient donor delivery systems are in vitro synthetic DNA and viral vectors 4,5 . Synthetic DNA donors are delivered to cells directly via electroporation or by packaging them into particles without specifically targeting the nucleus, while viral vectors such as adenoassociated virus (AAV) are transduced to enter the nucleus [6][7][8] .…”

Section: Introductionmentioning

confidence: 99%

Bacterial retrons enable precise gene editing in human cells

Zhao¹,

Chen²,

Lee³

et al. 2021

Preprint

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Retrons are bacterial genetic elements involved in anti-phage defense. They have the unique ability to reverse transcribe RNA into multicopy single-stranded DNA (msDNA) that remains covalently linked to their template RNA. Retrons coupled with CRISPR-Cas9 in yeast have been shown to improve editing efficiency of precise genome editing via homology-directed repair (HDR). HDR editing efficiency has been limited by challenges associated with delivering extracellular donor DNA encoding the desired mutation. In this study, we tested the ability of retrons to produce msDNA as donor DNA and facilitate HDR by tethering msDNA to guide RNA in HEK293T and K562 cells. Through heterologous reconstitution of retrons from multiple bacterial species with the CRISPR-Cas9 system, we demonstrated HDR rates of up to 11.3%. Overall, our findings represent the first step in extending retron-based precise gene editing to human cells.

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Recent Advances in CRISPR/Cas9 Delivery Strategies

Cited by 228 publications

References 103 publications

Various Aspects of a Gene Editing System—CRISPR–Cas9

Various Aspects of a Gene Editing System—CRISPR–Cas9

DNA-free gene editing in plants: a brief overview

Bacterial retrons enable precise gene editing in human cells

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