• Journal of Food Hygiene and Safety
• /
• v.38 no.5
• /
• pp.279-286
• /
• 2023

Rapid and accurate detection of pathogenic bacteria is crucial for various applications, including public health and food safety. However, existing bacteria detection techniques have several drawbacks as they are inconvenient and require time-consuming procedures and complex machinery. Recently, the precision and versatility of CRISPR/Cas system has been leveraged to design biosensors that offer a more efficient and accurate approach to bacterial detection compared to the existing techniques. Significant research has been focused on developing biosensors based on the CRISPR/Cas system which has shown promise in efficiently detecting pathogenic bacteria or virus. In this review, we present a biosensor based on the CRISPR/Cas system that has been specifically developed to overcome these limitations and detect different pathogenic bacteria effectively including Vibrio parahaemolyticus, Salmonella, E. coli O157:H7, and Listeria monocytogenes. This biosensor takes advantage of the CRISPR/Cas system's precision and versatility for more efficiently accurately detecting bacteria compared to the previous techniques. The biosensor has potential to enhance public health and ensure food safety as the biosensor's design can revolutionize method of detecting pathogenic bacteria. It provides a rapid and reliable method for identifying harmful bacteria and it can aid in early intervention and preventive measures, mitigating the risk of bacterial outbreaks and their associated consequences. Further research and development in this area will lead to development of even more advanced biosensors capable of detecting an even broader range of bacterial pathogens, thereby significantly benefiting various industries and helping in safeguard human health

• BMB Reports
• /
• v.54 no.1
• /
• pp.59-69
• /
• 2021

The ability to read, write, and edit genomic information in living organisms can have a profound impact on research, health, economic, and environmental issues. The CRISPR/Cas system, recently discovered as an adaptive immune system in prokaryotes, has revolutionized the ease and throughput of genome editing in mammalian cells and has proved itself indispensable to the engineering of immune cells and identification of novel immune mechanisms. In this review, we summarize the CRISPR/Cas9 system and the history of its discovery and optimization. We then focus on engineering T cells and other types of immune cells, with emphasis on therapeutic applications. Last, we describe the different modifications of Cas9 and their recent applications in the genome-wide screening of immune cells.

• Molecules and Cells
• /
• v.46 no.1
• /
• pp.4-9
• /
• 2023

The advent of the CRISPR-Cas genome editing platform has greatly enhanced the capabilities of researchers in many areas of biology. Its use has also been turned to the development of therapies for genetic diseases and to the enhancement of cell therapies. This review describes some recent advances in these areas.

• Molecules and Cells
• /
• v.41 no.10
• /
• pp.917-922
• /
• 2018

The CRISPR-Cas system is a well-established RNA-guided DNA editing technique widely used to modify genomic DNA sequences. I used the CRISPR-Cas9 system to change the second and third nucleotides of the triplet $T{\underline{CT}}$ of human TNSFSF9 in HepG2 cells to $T{\underline{AG}}$ to create an amber stop codon. The $T{\underline{CT}}$ triplet is the codon for Ser at the $172^{nd}$ position of TNSFSF9. The two substituted nucleotides, AG, were confirmed by DNA sequencing of the PCR product followed by PCR amplification of the genomic TNFSF9 gene. Interestingly, sequencing of the cDNA of transcripts of the edited TNFSF9 gene revealed that the $T{\underline{AG}}$ had been re-edited to the wild type triplet $T{\underline{CT}}$, and 1 or 2 bases just before the triplet had been deleted. These observations indicate that CRISPR-Cas9-mediated editing of bases in target genomic DNA can be followed by spontaneous re-editing (correcting) of the bases during transcription.

• BMB Reports
• /
• v.50 no.5
• /
• pp.247-256
• /
• 2017

CRISPR/Cas9 is the latest tool introduced in the field of genome engineering and is so far the best genome-editing tool as compared to its precedents such as, meganucleases, zinc finger nucleases (ZFNs) and transcription activator-like effectors (TALENs). The simple design and assembly of the CRISPR/Cas9 system makes genome editing easy to perform as it uses small guide RNAs that correspond to their DNA targets for high efficiency editing. This has helped open the doors for multiplexible genome targeting in many species that were intractable using old genetic perturbation techniques. Currently, The CRISPR system is revolutionizing the way biological researches are conducted and paves a bright future not only in research but also in medicine and biotechnology. In this review, we evaluated the history, types and structure, the mechanism of action of CRISPR/Cas System. In particular, we focused on the application of this powerful tool in autophagy research.

• Proceedings of the Korean Society of Crop Science Conference
• /
• 2019.04a
• /
• pp.9-9
• /
• 2019

• Journal of Ginseng Research
• /
• v.46 no.4
• /
• pp.505-514
• /
• 2022

Background: The roots of Panax ginseng contain two types of tetracyclic triterpenoid saponins, namely, protopanaxadiol (PPD)-type saponins and protopanaxatiol (PPT)-type saponins. In P. ginseng, the protopanaxadiol 6-hydroxylase (PPT synthase) enzyme catalyses protopanaxatriol (PPT) production from protopanaxadiol (PPD). In this study, we constructed homozygous mutant lines of ginseng by CRISPR/Cas9-mediated mutagenesis of the PPT synthase gene and obtained the mutant ginseng root lines having complete depletion of the PPT-type ginsenosides. Methods: Two sgRNAs (single guide RNAs) were designed for target mutations in the exon sequences of the two PPT synthase genes (both PPTa and PPTg sequences) with the CRISPR/Cas9 system. Transgenic ginseng roots were generated through Agrobacterium-mediated transformation. The mutant lines were screened by ginsenoside analysis and DNA sequencing. Result: Ginsenoside analysis revealed the complete depletion of PPT-type ginsenosides in three putative mutant lines (Cr4, Cr7, and Cr14). The reduction of PPT-type ginsenosides in mutant lines led to increased accumulation of PPD-type ginsenosides. The gene editing in the selected mutant lines was confirmed by targeted deep sequencing. Conclusion: We have established the genome editing protocol by CRISPR/Cas9 system in P. ginseng and demonstrated the mutated roots producing only PPD-type ginsenosides by depleting PPT-type ginsenosides. Because the pharmacological activity of PPD-group ginsenosides is significantly different from that of PPT-group ginsenosides, the new type of ginseng mutant producing only PPD-group ginsenosides may have new pharmacological characteristics compared to wild-type ginseng. This is the first report to generate target-induced mutations for the modification of saponin biosynthesis in Panax species using CRISPR-Cas9 system.

• Proceedings of the Korean Society of Crop Science Conference
• /
• 2022.10a
• /
• pp.288-288
• /
• 2022

Rice is an important staple in the world. And drought is one of the important constraints that negatively affect yield loss and grain quality of rice. CRISPR/Cas9 is a new breeding strategy that can improve the characteristics of rice quickly and accurately. CRISPR/Cas9 is a novel approach that can reliably harvest rice yields in response to a rapidly changing climate. In addition, there is no externally inserted DNA left in genome-editing rice, and it is receiving attention as being able to take responsibility for future food because its characteristics are continuously improved. In the future, high levels of drought resistant in water-constrained environments will be required, which will reduce yield loss. OsSAP was genome-editing with CRISPR/Cas9 in rice. A different line number was assigned to each panicle, and the generation advanced by applying the ear-to-row method. Genome-editing rice has improved drought resistance in drought conditions. Also, in genome-editing rice, the target sequence was homozygous in the 0 generation, and the coefficient of variation of heading date, number of tiller, and 1,000-grain weight was very small in 2 generation. In the era of rapidly changing climate change, CRISPR/Cas9 presents a new breeding strategy that can rapidly and accurately improve agronomic traits of major food crops as well as rice. CRISPR/Cas9 is applied together with traditional breeding to develop into a new breeding strategy, it is suggested that food can be obtained stably in response to climate change.

• Biomedical Science Letters
• /
• v.25 no.4
• /
• pp.372-380
• /
• 2019

Mice with specific modified genes are useful means of studying development and disease. The CRISPR/Cas9 system is a very powerful and effective tool for generating genetically modified mice in a simple and fast manner. To generate human IgG light kappa constant knock-in mice, we tested by microinjection of a mixture of Cas9 protein, single-guide RNA and target homologous recombinant donor DNA into zygotes. We found that the injection of 10 ng/μL of Cas9 protein and crRNA/tracrRNA, rather than single guide RNA, induced the production of knock-in mice more effectively. Thus, our study provides valuable information that will help to improve the production of knock-in mice and contribute the successful generation of humanized Ab-producing mice in Korea.

• Korean Journal of Agricultural Science
• /
• v.46 no.4
• /
• pp.885-895
• /
• 2019

Plant biotechnologists have long dreamed of technologies to manipulate genes in plants at will. This dream has come true partly through the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology, which now has been used to edit genes in several important crops. However, there are many restrictions in editing a gene precisely using the CRISPR/Cas9 technology because CRISPR/Cas9 may cause deletions or additions in some regions of the target gene. Several other technologies have been developed for gene targeting and precision editing. Among these, base editors might be the most practically and efficiently used compared to others. Base editors are tools which are able to cause a transition from cytosine into thymine, or from adenine into guanine very precisely on specific sequences. Cytosine base editors basically consist of nCas9, cytosine deaminase, and uracil DNA glycosylase inhibitor (UGI). Adenine base editors consist of nCas9 and adenine deaminase. These were first developed for human cells and have since also been applied successfully to crops. Base editors have been successfully applied for productivity improvement, fortification and herbicide resistance of crops. Thus, base editor technologies start to open a new era for precision gene editing or breeding in crops and might result in revolutionary changes in crop breeding and biotechnology.