• Journal of Plant Biotechnology
• /
• v.44 no.2
• /
• pp.107-114
• /
• 2017

Implementation of crop improvement programs relies on genetic diversity. To overcome the limited occurrence of natural mutations, researchers and breeders applied diverse methods, ranging from conventional crossing to classical bio-technologies. Earlier generations of knockout and gain-of-function technologies often result in incomplete gene disruption or random insertions of transgenes into plant genomes. The newly developed editing tool, CRISPR/Cas9 system, not only provides a powerful platform to efficiently modify target traits, but also broadens the scope and prospects of genome editing. Customized Cas9/guide RNA (gRNA) systems suitable for efficient genomic modification of mammalian cells or plants have been reported. Following successful demonstration of this technology in mammalian cells, CRISPR/Cas9 was successfully adapted in plants, and accumulating evidence of its feasibility has been reported in model plants and major crops. Recently, a modified version of CRISPR/Cas9 with added novel functions has been developed that enables programmable direct irreversible conversion of a target DNA base. In this review, we summarized the milestone applications of CRISPR/Cas9 in plants with a focus on major crops. We also present the implications of an improved version of this technology in the current plant breeding programs.

• Korean Journal of Agricultural Science
• /
• v.47 no.4
• /
• pp.903-920
• /
• 2020

Since its first demonstration as a practical genome editing tool in the early 2010s, the use of clustered regularly interspaced short palindromic repeat (CRISPR) along with the endonuclease Cas9 (CRISPR/Cas9) has become an essential choice for generating targeted mutations. Due to its relative simplicity and cost-effectiveness compared to other molecular scissors, i.e., zinc finger nuclease (ZFN) and transcription activator-like effector nuclease (TALEN), the CRISPR/Cas9 system has been shown to have a massive influence on genetic studies regardless of the biological kingdom. Although the system is in the process of being established, numerous protocols have already been released for the system and there have been various topics of CRISPR related papers published each year in ever-increasing manner. Here, we will briefly introduce CRISPR/Cas9 system and discuss the variants of the CRISPR system. Also, their applications to crop improvement will be dealt with mainly ornamental crops among horticultural crops other than Arabidopsis as a model plant. Finally, some issues on the barriers restraining the use of CRISPR system on floricultural crops, the prospect of CRISPR system as a DNA-free genome editing tool with efficient facilitators and finally, the future perspectives on the CRISPR system will be described.

• Journal of Life Science
• /
• v.34 no.8
• /
• pp.567-575
• /
• 2024

New breeding techniques (NBT) recognize specific DNA sequences and remove, modify, or insert DNA at a desired location, and can be used to treat genetic diseases in humans or to improve the traits of livestock or crops. In this study, we conducted a comparative analysis of various agricultural traits and assessed the safety of gene transferability in third-generation genome-editing rice (OsCKq1-G₃) with T and G nucleotide insertions developed using the CRISPR/Cas9 SDN-1 method, in comparison to its parental line (Oryza sativa L., cv Ilmi). The analyzed traits included heading date, culm length, panicle length, tiller number, yield, germination rate, viviparous germination rate, shattering, after wintering seed viability, the presence of toxins and allergens. The target trait, heading date, exhibited a high significant difference of approximately 5 days. Culm length, panicle length, tiller number, yield showed no significant differences compared to the parental line. No T-DNA bands indicating gene transfer were detected. In the third generation of genome-edited rice, the T-DNA was confirmed to be eliminated as successive generations advanced through self-pollination. Through the analysis of germination rate, viviparous germination rate, shattering, and after wintering viability, we confirmed that the genome-editing rice has no potential for weediness. The ORF and amino acid sequences of the genome-editing rice did not reveal any toxins and allergens. The results of this study can be utilized as important data for the environmental risk assessment of genome-editing rice.

• Journal of Plant Biotechnology
• /
• v.47 no.2
• /
• pp.107-117
• /
• 2020

Flower industry is now growing due to the development of economy in many countries. Simultaneously, needs from consumers in flower market are varied widely. To satisfy the needs from consumers and deal with a variety of diseases from a lots of pathogens as well as climate change, new elite flower cultivars should be released in flower market. For this purpose, conventional and biotechnological techniques can be employed to make good cultivar. Therefore, this review describes the general overview of flower breeding techniques including cross-hybridization, mutation breeding and genetic transformation systems. Also, breeding systems for ornamentals derived from plant tissue culture such as embryo culture, in vitro fertilization, ovary/ovule culture and haploid production were reviewed. Furthermore, in this study recent development of the generation of new flower cultivars using marker-assisted breeding, plant transformation including particle bombardment and Agrobacterium tumefaciens as well as genome-editing technology were described. This review will be contributed to the development and releasement of new flower cultivars with horticulturally useful traits in the future.

• Molecules and Cells
• /
• v.41 no.10
• /
• pp.889-899
• /
• 2018

Intrinsically disordered proteins (IDPs) are highly unorthodox proteins that do not form three-dimensional structures under physiological conditions. The discovery of IDPs has destroyed the classical structure-function paradigm in protein science, 3-D structure = function, because IDPs even without well-folded 3-D structures are still capable of performing important biological functions and furthermore are associated with fatal diseases such as cancers, neurodegenerative diseases and viral pandemics. Pre-structured motifs (PreSMos) refer to transient local secondary structural elements present in the target-unbound state of IDPs. During the last two decades PreSMos have been steadily acknowledged as the critical determinants for target binding in dozens of IDPs. To date, the PreSMo concept provides the most convincing structural rationale explaining the IDP-target binding behavior at an atomic resolution. Here we present a brief developmental history of PreSMos and describe their common characteristics. We also provide a list of newly discovered PreSMos along with their functional relevance.

• BMB Reports
• /
• v.57 no.1
• /
• pp.1-1
• /
• 2024

• BMB Reports
• /
• v.49 no.9
• /
• pp.497-501
• /
• 2016

Wide-line ¹H NMR intensity and differential scanning calorimetry measurements were carried out on the intrinsically disordered 73-residue full transactivation domain (TAD) of the p53 tumor suppressor protein and two peptides: one a wild type p53 TAD peptide with a helix pre-structuring property, and a mutant peptide with a disabled helix-forming propensity. Measurements were carried out in order to characterize their water and ion binding characteristics. By quantifying the number of hydrate water molecules, we provide a microscopic description for the interactions of water with a wild-type p53 TAD and two p53 TAD peptides. The results provide direct evidence that intrinsically disordered proteins (IDPs) and a less structured peptide not only have a higher hydration capacity than globular proteins, but are also able to bind a larger amount of charged solute ions.

• Biomedical Science Letters
• /
• v.26 no.2
• /
• pp.114-119
• /
• 2020

Of the various types of mice used for genome editing, conditional knock-out (cKO) mice serve as an important model for studying the function of genes. cKO mice can be produced using loxP knock-in (KI) mice in which loxP sequences (34 bp) are inserted on both sides of a specific region in the target gene. These mice can be used as KO mice that do not express a gene at a desired time or under a desired condition by cross-breeding with various Cre Tg mice. Genome editing has been recently made easy by the use of third-generation gene scissors, the CRISPR-Cas9 system. However, very few laboratories can produce mice for genome editing. Here we present a more efficient method for producing loxP KI mice. This method involves the use of an HDR vector as the target vector and ssODN as the donor DNA in order to induce homologous recombination for producing loxP KI mice. On injecting 20 ng/µL of ssODN, it was observed that the target exon was deleted or loxP was inserted on only one side. However, on injecting 10 ng/µL of the target HDR vector, the insertion of loxP was observed on both sides of the target region. In the first PCR, seven mice were identified to be loxP KI mice. The accuracy of their gene sequences was confirmed through Sanger sequencing. It is expected that the loxP KI mice produced in this study will serve as an important tool for identifying the function of the target gene.

• Molecules and Cells
• /
• v.42 no.8
• /
• pp.604-616
• /
• 2019

Phosphoserine phosphatase (PSPH) is one of the key enzymes of the L-serine synthesis pathway. PSPH is reported to affect the progression and survival of several cancers in an L-serine synthesis-independent manner, but the mechanism remains elusive. We demonstrate that PSPH promotes lung cancer progression through a noncanonical L-serine-independent pathway. PSPH was significantly associated with the prognosis of lung cancer patients and regulated the invasion and colony formation of lung cancer cells. Interestingly, L-serine had no effect on the altered invasion and colony formation by PSPH. Upon measuring the phosphatase activity of PSPH on a serine-phosphorylated peptide, we found that PSPH dephosphorylated phospho-serine in peptide sequences. To identify the target proteins of PSPH, we analyzed the protein phosphorylation profile and the PSPH-interacting protein profile using proteomic analyses and found one putative target protein, IRS-1. Immunoprecipitation and immunoblot assays validated a specific interaction between PSPH and IRS-1 and the dephosphorylation of phospho-IRS-1 by PSPH in lung cancer cells. We suggest that the specific interaction and dephosphorylation activity of PSPH have novel therapeutic potential for lung cancer treatment, while the metabolic activity of PSPH, as a therapeutic target, is controversial.

• International Journal of Industrial Entomology and Biomaterials
• /
• v.39 no.1
• /
• pp.14-21
• /
• 2019

Genome editing by CRISPR/Cas9, a third-generation gene scissor in molecular breeding at the genome level, is attracting much attention as one of the breeding techniques of the future. In this study, genetic and phenotypic analysis was used to examine the responsiveness of the Bakokjam variety of the silkworm Bombyx mori to molecular breeding using CRISPR/Cas9 in editing the white egg 2 (w-2) gene. The nucleotide sequence of the w-2 gene was analyzed and three different guide RNAs (gRNA) were prepared. The synthesized gRNA was combined with Cas9 protein and then analyzed by T7 endonuclease I after introduction into the Bm-N silkworm cell line. To edit the silkworm gene, W1N and W2P gRNA and Cas9 complexes were microinjected into silkworm embryos. Based on the results of microinjection, the hatching rate was 16-24% and the incidence of mutation was 33-37%. The gene mutation was verified in the heterozygous F1 generation, but no phenotypic change was observed. In F2 homozygotes generated by F1 self-crosses, a mutant phenotype was observed. These results suggest that silkworm molecular breeding using the CRISPR/Cas9 system is possible and will be a very effective way to shorten the time required than the traditional breeding process.