• Journal of Life Science
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• v.29 no.5
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• pp.537-544
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• 2019

Genome editing technology employing gene scissors has generated interest in molecular breeding in various fields, and the development of the third-generation gene scissors of the clustered, regularly interspaced short palindromic repeat (CRISPR) system has accelerated the field of molecular breeding through genome editing. In this study, we analyzed the possibility of silkworm molecular breeding using gene scissors by genomic and phenotypic analysis after editing the biogenesis of lysosome-related organelles (BmBLOS) gene of Bakokjam using the CRISPR/Cas9 system. Three types of guide RNAs (gRNA) were synthesized based on the BmBLOS gene sequence of Bakokjam. Complexes of the prepared gRNA and Cas9 protein were formed and introduced into Bombyx mori BM-N cells by electroporation. Analysis of the gene editing efficiency by T7 endonuclease I analysis revealed that the B4N gRNA showed the best efficiency. The silkworm genome was edited by microinjecting the Cas9/B4N gRNA complex into silkworm early embryos and raising the silkworms after hatching. The hatching rate was as low as 18%, but the incidence of mutation was over 40%. In addition, phenotypic changes were observed in about 70% of the G0 generation silkworms. Sequence analysis showed that the BmBLOS gene appeared to be a heterozygote carrying the wild-type and mutation in most individuals, and the genotype of the BmBLOS gene was also different in all individuals. These results suggest that although the possibility of silkworm molecular breeding using the CRISPR/Cas9 system would be very high, continued research on breeding and screening methods will be necessary to improve gene editing efficiency and to obtain homozygotes.

• Microbiology and Biotechnology Letters
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• v.49 no.4
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• pp.534-542
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• 2021

Single-molecular guide RNA (sgRNA) plays a role in recognizing the DNA target sequence in CRISPR technology for genome editing and gene expression control. In this study, we systematically compared the length of the target recognition sequence in sgRNAs required for genome editing using Cas9-NG (an engineered Cas9 recognizing 5'-NG as PAM sequence) and gene expression control using deactivated Cas9-NG (dCas9-NG) by targeting the gal promoter in E. coli. In the case of genome editing, the truncation of three nucleotides in the target recognition sequence (TRS) of sgRNA was allowed. In gene expression regulation, we observed that target recognition and binding were possible even if eleven nucleotides were deleted from twenty nucleotides of the TRS. When 4 or more nucleotides are truncated in the TRS of the sgRNA, it is thought that the sgRNA/Cas9-NG complex can specifically bind to the target DNA sequence, but lacks endonuclease activity to perform genome editing. Our study will be helpful in the development of artificial transcription factors and various CRISPR technologies in the field of synthetic biology.

• Journal of Veterinary Science
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• v.21 no.2
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• pp.26.1-26.14
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• 2020

Pancreatic ductal adenocarcinoma is a lethal cancer type that is associated with multiple gene mutations in somatic cells. Genetically engineered mouse is hardly applicable for developing a pancreatic cancer model, and the xenograft model poses a limitation in the reflection of early stage pancreatic cancer. Thus, in vivo somatic cell gene engineering with clustered regularly interspaced short palindromic repeats is drawing increasing attention for generating an animal model of pancreatic cancer. In this study, we selected Kras, Trp53, Ink4a, Smad4, and Brca2 as target genes, and applied Campylobacter jejuni Cas9 (CjCas9) and Streptococcus pyogens Cas9 (SpCas9) for developing pancreatic cancer using adeno associated virus (AAV) transduction. After confirming multifocal and diffuse transduction of AAV2, we generated SpCas9 overexpression mice, which exhibited high double-strand DNA breakage (DSB) in target genes and pancreatic intraepithelial neoplasia (PanIN) lesions with two AAV transductions; however, wild-type (WT) mice with three AAV transductions did not develop PanIN. Furthermore, small-sized Cjcas9 was applied to WT mice with two AAV system, which, in addition, developed high extensive DSB and PanIN lesions. Histological changes and expression of cancer markers such as Ki67, cytokeratin, Mucin5a, alpha smooth muscle actin in duct and islet cells were observed. In addition, the study revealed several findings such as 1) multiple DSB potential of AAV-CjCas9, 2) peri-ductal lymphocyte infiltration, 3) multi-focal cancer marker expression, and 4) requirement of > 12 months for initiation of PanIN in AAV mediated targeting. In this study, we present a useful tool for in vivo cancer modeling that would be applicable for other disease models as well.

• Publications of The Korean Astronomical Society
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• v.20 no.1 s.24
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• pp.29-35
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• 2005

Time-series observations were carried out using a 155mm refractor and a $2k{\times}3k$ CCD camera at Bohyunsan Optical Astronomy Observatory. We found 38 new variable stars in the $2.3^{\circ}{\times}2.4^{\circ}$ region around the Cepheid variable TU Cas: 22 eclipsing binary stars, 11 ${\delta}$ Scuti type stars and an RR Lyrae star, and 4 unclassified variables.

• Journal for History of Mathematics
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• v.24 no.4
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• pp.157-180
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• 2011

This paper analyzed about gender difference in the achievement of underachievers of high school students while learning using technology. Participants were composed of 67 underachievers on first grade in high school located in a metropolitan city. That had never used a mathematics educational calculator before. Target participants were divided into two groups: experiment group that studied activity papers with a CAS calculator. And control group that studied the same activity papers using only paper-and-pencil. The content of the activity papers for the two groups was the same, but the structure differed. The two groups completed mathematics achievement tests both before and after the activity papers. The results are that find out no difference of the mathematics achievement between boys and girls in each group, and that the mathematics achievement of boys in experiment group are better than one of boys in control group, and also girls.

• Proceedings of the Korean Institute of Navigation and Port Research Conference
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• 2019.11a
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• pp.264-264
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• 2019

The purpose of this paper is to establish development concepts for a novel collision avoidance system with preventing function of navigator's human error (Hu-CAS) in ship control behaviors. Hu-CAS consists of four modules: 1) collision risk assessment module to estimate collision priority between the ship and objects, 2) decision-making module to decide collision risk levels, 3) parameter estimation module needed in the ship control to avoid collisions and 4) control system to control the rudder angle and speed. Hu-CAS, proposed in this paper, can provide a novel system substitution current Autopilot and/or a CAS be teen manned vessel and Autonomous ship in a future.

• Development and Reproduction
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• v.27 no.4
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• pp.205-211
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• 2023

INTS14/VWA9, a component of the integrator complex subunits, plays a pivotal role in regulating the fate of numerous nascent RNAs transcribed by RNA polymerase II, particularly in the biogenesis of small nuclear RNAs and enhancer RNAs. Despite its significance, a comprehensive mutation model for developmental research has been lacking. To address this gap, we aimed to investigate the expression patterns of INTS14 during zebrafish embryonic development. We generated ints14 mutant strains using the CRISPR/Cas9 system. We validated the gRNA activity by co-injecting Cas9 protein and a single guide RNA into fertilized zebrafish eggs, subsequently confirming the presence of a 6- or 9-bp deletion in the ints14 gene. In addition, we examined the two mutant alleles through PCR analysis, T7E1 assay, TA-cloning, and sequencing. For the first time, we used the CRISPR/Cas9 system to create a model in which some sequences of the ints14 gene were removed. This breakthrough opens new avenues for in-depth exploration of the role of ints14 in animal diseases. The mutant strains generated in this study can provide a valuable resource for further investigations into the specific consequences of ints14 gene deletion during zebrafish development. This research establishes a foundation for future studies exploring the molecular mechanisms underlying the functions of ints14, its interactions with other genes or proteins, and its broader implications for biological processes.

• Journal of the Korean Society of Food Science and Nutrition
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• v.41 no.8
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• pp.1168-1175
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• 2012

Gel properties of Mook manufactured from acorn harvested in various countries [domestic (KAS), Chinese (CAS), and North Korea (NAS)] were analyzed according to storage period. Gel properties included water binding, syneresis of gel, DSC (differential scanning calorimetry), X-ray diffraction, SEM (scanning electron microscopy), texture, Hunter's color value, and sensory value. Water binding of NAS, CAS, and KAS were 233.8%, 217.3%, and 215.0%, respectively. Syneresis of gel from KAS, CAS, and NAS were 2.06%, 1.85%, and 1.45%, respectively, after 1 day of storage. There were significant differences upon storage for 1~3 days (p<0.05), whereas were no significant differences upon storage for 4~10 days. Peak temperature of gelatinization property by DSC was $55.28^{\circ}C$ for KAS, $54.45^{\circ}C$ for CAS, and $54.12^{\circ}C$ for NAS after 1 day of storage. Hardness of texture in NAS, KAS, and CAS were 374.9, 357.4, and 348.9, respectively, after 1 day of storage. Hunter's color L value, and a value were highest in NAS, whereas b value was the lowest in CAS. There were no significant differences in any particular sensory values.

• Journal of Korean Society of Environmental Engineers
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• v.39 no.4
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• pp.201-207
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• 2017

This study was conducted to investigate the characteristics of the stream current detector (SCD) and charge analyzing system (CAS), which are well known as the automatic coagulant dosage determination instruments. When the SCD system was used, the current charge value set at pH 9.3 could not be used as the coagulant dosage when the pH of the raw water decreased to 5.7, and the current charge value corresponding to pH 5.7 was required to be reset. In case of CAS system, polydadmac is stable as a cationic titrant when the pH of the raw water ranged 9.3 to 6.0. And it was effective to use polydadmac as a cationic titrant for high turbidity water. The current charge of polydadmac was almost constant between pH 5 and 11, however, the difference in charge of alum is six times in this pH range. Therefore, when the pH of the raw water was changed, it seems that the coagulant used in the actual water treatment plant was as a good cationic titrant.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.7
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• pp.1029-1036
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• 2017

Objective: In the livestock industry, the regulatory mechanisms of muscle proliferation and differentiation can be applied to improve traits such as growth and meat production. We investigated the regulatory pathway of MyoD and its role in muscle differentiation in quail myoblast cells. Methods: The MyoD gene was mutated by the clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 technology and single cell-derived MyoD mutant sublines were identified to investigate the global regulatory mechanism responsible for muscle differentiation. Results: The mutation efficiency was 73.3% in the mixed population, and from this population we were able to establish two QM7 MyoD knockout subline (MyoD KO QM7#4) through single cell pick-up and expansion. In the undifferentiated condition, paired box 7 expression in MyoD KO QM7#4 cells was not significantly different from regular QM7 (rQM7) cells. During differentiation, however, myotube formation was dramatically repressed in MyoD KO QM7#4 cells. Moreover, myogenic differentiation-specific transcripts and proteins were not expressed in MyoD KO QM7#4 cells even after an extended differentiation period. These results indicate that MyoD is critical for muscle differentiation. Furthermore, we analyzed the global regulatory interactions by RNA sequencing during muscle differentiation. Conclusion: With CRISPR/Cas9-mediated genomic editing, single cell-derived sublines with a specific knockout gene can be adapted to various aspects of basic research as well as in functional genomics studies.