• Journal of Plant Biotechnology
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• v.44 no.1
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• pp.89-96
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• 2017

The CRISPR/Cas9 is a core technology that can result in a paradigm for breeding new varieties. This study describes in detail the sgRNA design, vector construction, and the development of a transgenic plant and its molecular analysis, and demonstrates how gene editing technology through the CRISPR/Cas9 system can be applied easily and accurately. CRISPR/Cas9 facilitates targeted gene editing through RNA-guided DNA cleavage, followed by cellular DNA repair mechanisms that introduce sequence changes at the site of cleavage. It also allows the generation of heritable-targeted gene mutations and corrections. Here, we present detailed procedures involved in the CRISPR/Cas9 system to acquire faster, easier and more cost-efficient gene edited transgenic rice. The protocol described here establishes the strategies and steps for the selection of targets, design of sgRNA, vector construction, and analysis of the transgenic lines. The same principles can be used to customize the versatile CRISPR/Cas9 system, for application to other plant species.

• Molecules and Cells
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• v.37 no.7
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• pp.547-553
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• 2014

Glioblastoma multiforme (GBM) is one of the most common brain malignancies and has a very poor prognosis. Recent evidence suggests that the presence of cancer stem cells (CSC) in GBM and the rare CSC subpopulation that is resistant to chemotherapy may be responsible for the treatment failure and unfavorable prognosis of GBM. A garlic-derived compound, Z-ajoene, has shown a range of biological activities, including anti-proliferative effects on several cancers. Here, we demonstrated for the first time that Z-ajoene specifically inhibits the growth of the GBM CSC population. CSC sphere-forming inhibition was achieved at a concentration that did not exhibit a cytotoxic effect in regular cell culture conditions. The specificity of this inhibitory effect on the CSC population was confirmed by detecting CSC cell surface marker CD133 expression and biochemical marker ALDH activity. In addition, stem cell-related mRNA profiling and real-time PCR revealed the differential expression of CSC-specific genes, including Notch, Wnt, and Hedgehog, upon treatment with Z-ajoene. A proteomic approach, i.e., reverse-phase protein array (RPPA) and Western blot analysis, showed decreased SMAD4, p-AKT, 14.3.3 and FOXO3A expression. The protein interaction map (http://string-db.org/) of the identified molecules suggested that the AKT, ERK/p38 and $TGF{\beta}$ signaling pathways are key mediators of Z-ajoene's action, which affects the transcriptional network that includes FOXO3A. These biological and bioinformatic analyses collectively demonstrate that Z-ajoene is a potential candidate for the treatment of GBM by specifically targeting GBM CSCs. We also show how this systemic approach strengthens the identification of new therapeutic agents that target CSCs.

• Journal of Animal Science and Technology
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• v.46 no.4
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• pp.525-536
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• 2004

The objective of this study was to identify the quantitative traits loci(QTL) for meat quality traits in pigs. Three-generation resource population was constructed from a cross between Korean native boars and Landrace sows. The resource population including founders, $F_1$ and $F_2$ was genotyped for 23 microsatellite markers on chromosome 7. The sex average total length of linkage map on chromosome 7 was estimated 154.6 cM. Meat quality traits including meat pH, meat color, drip loss, shear force, heating loss, crude fat, crude protein, crude ash and water content in muscle were collected from $F_2$ animals. For the QTL mapping, we used $F_2$ QTL Analysis Servlet of QTL express for web-based QTL mapping tools(http://qtl.cap.ed.ac.uk/). The QTLs for CIE-a and CIE-b on SSC7 were significantly detected at 1% and 5% chromosome-wide level, respectively.

• Journal of Ginseng Research
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• v.38 no.4
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• pp.278-288
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• 2014

Background: Panax ginseng Meyer is a traditional medicinal plant famous for its strong therapeutic effects and serves as an important herbal medicine. To understand and manipulate genes involved in secondary metabolic pathways including ginsenosides, transcriptome profiling of P. ginseng is essential. Methods: RNA-seq analysis of adventitious roots of two P. ginseng cultivars, Chunpoong (CP) and Cheongsun (CS), was performed using the Illumina HiSeq platform. After transcripts were assembled, expression profiling was performed. Results: Assemblies were generated from ~85 million and ~77 million high-quality reads from CP and CS cultivars, respectively. A total of 35,527 and 27,716 transcripts were obtained from the CP and CS assemblies, respectively. Annotation of the transcriptomes showed that approximately 90% of the transcripts had significant matches in public databases.We identified several candidate genes involved in ginsenoside biosynthesis. In addition, a large number of transcripts (17%) with different gene ontology designations were uniquely detected in adventitious roots compared to normal ginseng roots. Conclusion: This study will provide a comprehensive insight into the transcriptome of ginseng adventitious roots, and a way for successful transcriptome analysis and profiling of resource plants with less genomic information. The transcriptome profiling data generated in this study are available in our newly created adventitious root transcriptome database (http://im-crop.snu.ac.kr/transdb/index.php) for public use.

• Journal of Animal Science and Technology
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• v.45 no.6
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• pp.939-948
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• 2003

The objective of this study was to identify the quantitative traits loci(QTL) for economically important traits such as growth, carcass and meat quality on pig chromosome 6. A three generation resource population was constructed from cross between Korean native boars and Landrace sows. A total of 240 F$_2$ animals were produced using intercross between 10 boars and 31 sows of F$_1$ animals. Phenotypic data including body weight at 3 weeks, backfat thickness, muscle pH, shear force and crude protein level were collected from F$_2$ animals. Animals including grandparents(F$_0$), parents(F$_1$) and offspring(F$_2$) were genotyped for 29 microsatellite markers and PCR-RFLP marker on chromosome 6. The linkage analysis was performed using CRI-MAP software version 2.4(Green et al., 1990) with FIXED option to obtain the map distances. The total length of SSC6 linkage map estimated in this study was 169.3cM. The average distance between adjacent markers was 6.05cM. For mapping of QTL, we used F$_2$ QTL Analysis Servlet of QTL express, a web-based QTL mapping tool(http://qtl.cap.ed.ac.uk). Five QTLs were detected at 5% chromosome-wide level for body weight of 3 weeks of age, shear force, meat pH at 24 hours after slaughtering, backfat thickness and crude protein level on SSC6.

• Journal of Life Science
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• v.16 no.3 s.76
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• pp.454-458
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• 2006

HSF1 is the heat shock transcription factor in Saccharomyces cerevisiae. S. cerevisiae KNU5377 can ferment at high temperature such as $40^{\b{o}}C$. We have been the subjects of intense study because Hsf1p mediates gene expression not only to heat shock, but to a variety of cellular and environmental stress challenges. Basing these facts, we firstly tried to construct the hsf1 gene-deleted mutant. PCR-method for fast production of gene disruption cassette was introduced in a thermotolerant yeast S. cerevisiae KNU5377, which allowed the addition of short flanking homology region as short as 45 bp suffice to mediate homologous recombination to kanMX module. Such a cassette is composed of linking genomic DNA of target gene to the selectable marker kanMX4 that confers geneticin (G418) resistance in yeast. That module is extensively used for PCR-based gene replacement of target gene in the laboratory strains. We describe here the generation of hsf1 gene disruption construction using PCR product of selectable marker with primers that provide homology to the hsf1 gene following separation of haploid strain in wild type yeast S. cerevisiae KNU5377. Yeast deletion overview containing replace cassette module, deletion mutant construction and strain confirmation in this study used Saccharomyces Genome Deletion Project (http:://www-sequence.standard.edu/group/yeast_deletion_project). This mutant by genetic manipulation of wild type yeast KNU5377 strain will provide a good system for analyzing the research of the molecular biology underlying their physiology and metabolic process under fermentation and improvement of their fermentative properties.

• BMB Reports
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• v.38 no.6
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• pp.739-747
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• 2005

Full-length cDNA sequences of four novel SPATA4 genes in chimpanzee, cow, chicken and ascidian were identified by bioinformatic analysis using mouse or human SPATA4 cDNA fragment as electronic probe. All these genes have 6 exons and have similar protein molecular weight and do not localize in sex chromosome. The mouse SPATA4 sequence is identified as significantly changed in cryptorchidism, which shares no significant homology with any known protein in swissprot databases except for the homologous genes in various vertebrates. Our searching results showed that all SPATA4 proteins have a putative conserved domain DUF1042. The percentages of putative SPATA4 protein sequence identity ranging from 30% to 99%. The high similarity was also found in 1 kb promoter regions of human, mouse and rat SPATA4 gene. The similarities of the sequences upstream of SPATA4 promoter also have a high proportion. The results of searching SymAtlas (http://symatlas.gnf.org/SymAtlas/) showed that human SPATA4 has a high expression in testis, especially in testis interstitial, leydig cell, seminiferous tubule and germ cell. Mouse SPATA4 was observed exclusively in adult mouse testis and almost no signal was detected in other tissues. The pI values of the protein are negative, ranging from 9.44 to 10.15. The subcellular location of the protein is usually in the nucleus. And the signal peptide possibilities for SPATA4 are always zero. Using the SNPs data in NCBI, we found 33 SNPs in human SPATA4 gene genomic DNA region, with the distribution of 29 SNPs in the introns. CpG island searching gives the data about CpG island, which shows that the regions of the CpG island have a high similarity with each other, though the length of the CpG island is different from each other.This research is a fundamental work in the fields of the bioinformational analysis, and also put forward a new way for the bioinformatic analysis of other genes.

• Proceedings of the Korean Society of Crop Science Conference
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• 2017.06a
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• pp.140-140
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• 2017

Nutritious and functional foods from crop have received great attention in recent years. Colored-grain wheat contains high phenolic compound and a large number of flavonoid. The anthocyanin and polyphenolic synthesis and accumulation is generally stimulated in response to biotic or abiotic stresses. Here, we analyzed genome wide transcripts in seedling of colored-grain wheat response to ABA and PEG treatment. About 900 and 1500 transcripts (p-value < 0.05) from ABA and PEG treatment were aligned to IWGSC1+popseq DB which is composed of over 110,000 transcripts including 100,934 coding genes. NR protein sequences of Poaceae from NCBI and protein sequence of transcription factors originated from 83 species in plant transcription factor database v3.0 were used for annotation of putative transcripts. Gene ontology analysis were conducted and KEGG mapping was performed to show expression pattern of biosynthesis genes related in flavonoid, isoflavonoid, flavons and anthocyanin biopathway. DroughtDB (http://pgsb.helmholtz-muenchen.de/droughtdb/) was used for detection of DEGs to explain that physiological and molecular drought avoidance by drought tolerance mechanisms. Drought response pathway, such as ABA signaling, water and ion channels, detoxification signaling, enzymes of osmolyte biosynthesis, phospholipid metabolism, signal transduction, and transcription factors related DEGs were selected to explain response mechanism under water deficit condition. Anthocyanin, phenol compound, and DPPH radical scavenging activity were measured and antioxidant activity enzyme assays were conducted to show biochemical adaptation under water deficit condition. Several MYB and bHLH transcription factors were up-regulated in both ABA and PEG treated condition, which means highly expressed MYB and bHLH transcription factors enhanced the expression of genes related in the biosynthesis pathways of flavonoids, such as anthocyanin and dihydroflavonols in colored wheat seedlings. Subsequently, the accumulation of total anthocyanin and phenol contents were observed in colored wheat seedlings, and antioxidant capacity was promoted by upregulation of genes involved in maintaining redox state and activation of antioxidant scavengers, such as CAT, APX, POD, and SOD in colored wheat seedlings under water deficit condition. This work may provide valuable and basic information for further investigation of the molecular responses of colored-grain wheat to water deficit stress and for further gene-based studies.

• Journal of Korean Medicine Rehabilitation
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• v.23 no.3
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• pp.117-124
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• 2013

Objectives The aims of this study was to observe the effectiveness of East-West Integrative Medicine (EWIM) by analysing between EWIM estimated group and Non-EWIM eastimated group with Brain Rehabilitation Registry. Methods We divided hospitals into two groups, one is EWIM estimated group hospital (Kyeonghee university, Wonkwang university, Dongguk university) and the other is non-EWIM estimated group (Jeonnam university, Gyeongbuk university, Catholic university). Then we analysed the effectiveness of treatments retrospectively using the database of Brain Rehabilitation Registry (http://www.kbrr.or.kr/, version 1.0). Totally 293 patients were included, EWIM estimated group was 175 and non-EWIM estimated group was 118. The main outcome measurements were National Institutes of Health Stroke Scale (NIHSS), Modified Barthel Index (MBI) and Mini Mental State Examination-Korea (MMSE-K). Results Changes of NIHSS was not significant in both total patient and patient who treated over 3 weeks. MMSE-K showed positively significant difference (p=0.044) in EWIM estimated group patients who treated over 3 weeks. In case of MBI, EWIM estimated group showed more effective result and also statistically significant in both total patients and patients who treated over 3 weeks. Conclusions We patially argued that EWIM estimated group was more effective than non-EWIM estimated group in stroke patients' functional recovery. We suggested base data of EWIM in stroke patiens through this study and this could be applied future researches of developing modified EWIM system.

• Proceedings of the Korea Water Resources Association Conference
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• 2015.05a
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• pp.441-441
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• 2015

북한의 수자원 현황을 분석하고 미래를 전망하는 일은 수자원 분야 전문가들의 공통된 관심사 중 하나이다. 그러나 많은 노력에도 불구하고 제한된 자료로 인해 이에 대한 체계적인 성과가 구축되지 못한 것이 사실이다(안재현 등, 2010). 북한 수자원 현황과 관련해서 1994년 한국수자원공사의"북한 수자원 현황 및 개발 동향"에 대한 연구와 1996년 한국과학기술단체총연합회의"남북한 수자원 비교평가 연구"가 수행된 바 있다. 본 연구에서는 2012년 7월 북한의 홍수피해에 대하여 북한지역의 빈약한 강우량 정보를 활용하여 위성으로부터 유도된 강우자료를 보정하여 북한지역의 홍수량을 산정하는데 활용할 수 있는 시계열 자료를 생성하는 것을 목적으로 하였다. 2012년 북한 홍수피해는 조선 중앙통신 보도에 의하면 6월 말부터 7월말까지 169명 사망, 400여명 실중, 144명 상해의 인명피해와 평안남북도, 자강도, 함경남북도 등 4만 3770여 가구가 침수되었으며, 농경지 피해는 6만 5280정보가 유실, 매몰, 침수(평안북도 2만 3400정보, 평안남도 2만 1900정보, 함경남도 5679정보, 함경북도 7220정보)의 피해를 입은 것으로 보도하고 있다(http://www.hani.co.kr/arti/politics/defense/545795.html). 한편, 북한의 기상수문국 통보에 의하면 2012년 장마가 시작된 이후 20일 동인 평균 강우량의 2배 가까운 비가 내렸으며, 특히 7월 19일 오후 9시부터 21일 오후 3시 사이 자강도 동신군에 413mm, 송원군에 383mm, 희천시에 322mm, 평안북도 태천군에 380mm, 향산군에 312mm, 동창군에 304mm의 폭우가 쏟아졌다고 조선중앙통신은 덧붙였다. 본 연구에서는 위성으로부터 유도된 강수자료 중 GSMaP_NRT자료를 활용하여 북한의 기상수문국에서 통보한 같이 기간의 강수량 자료를 분석한 결과 자강도 동신군 지역의 경우 총강우는 208.8mm로 분석되었으며, 평균강우는 9.49mm, 최대강우강도는 17.7mm/hr로 분석되었다. 두 자료의 총강우량에 대한 상대오차는 약 49.5% 정도의 오차가 있는 것으로 분석되었다. 위성강우 보정 방법으로 Immerzeel(2001) 등은 위성으로 관측된 시계열 자료와 지상계측된 시계열 자료를 분석하여 두 자료간의 평균과 표준편차를 이용하여 평균 및 표준편차 계수를 산정하여 위성자료에 대한 보정강우를 생성하는 방법을 제시한 바 있다. 본 연구에서는 위성으로부터 유도된 강우량 자료는 시계열 자료이나 지점계측 강우는 총강우만 존재하여 총강우에 대한 시계열 자료를 위성강수자료를 무차원화하여 지점계측의 위성 보정강우를 생성하였다.