• Bulletin of the Korean Chemical Society
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• 제34권4호
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• pp.1300-1302
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• 2013

• 미생물학회지
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• 제30권2호
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• pp.141-148
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• 1992

In order to overexpress bacteriophage PRD1 DNA polymerase in E. coli cells, the 2 kb HaeII fragment was isolated from phage genomic DNA. This fragment was then cloned into pEMBL/sup ex/ 3-expression vector. A specific 57bp deletion was performed by using uracil containing ss DNA and oligonucleotide spanning each region to remove an unwanted non-coding region. After this deletion, the PRD1 DNA polymerase gene is totally under the control of the vector promoter and SD sequence. Upon heat induction, a protein with an apparent size of 68 kdal was overexpressed as an active PRD1 DNA polymerase. The expression of PRD1 DNA polymerase was about 1% of total E. coli protein.

• 한국식물병리학회지
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• 제14권2호
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• pp.177-183
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• 1998

A DNA fragment which is involved in tolassin production was cloned to obtain a molecular marker of Pseudomonas tolaasii, a casual agent of bacterial brown blotch disease of oyster mushroom (Pleurotus ostreatus). Tolaasin is a lipodepsipeptide toxin and known as a primary disease determinant of the P. tolaasii. It is responsible for formation of white line in agar when P. tolaasii were cultured against white line reacting organisms (WLROs). White line negative mutants (WL-) were generated by conjugation between rifampicin resistant strain of P. tolaasii and E. coli carrying suicidal plasmid pSUP2021 : : Tn5. The ability of tolaasin production of the WL- mutants was examined by hemolysis test, pathogenicity test, and high pressure liquid chromatography (HPLC) analysis of culture filtrate. All of the WL- mutants were lost the ability of tolaasin production (Tol-). Genomic library of the Tol- mutant was constructed in pLAFR3 and the cosmid clone containing Tn5 was selected. DNA fragment fro franking region of Tn5 was cloned from the plasmid and used as a probe in Southern blot. DNA-DNA hybridization with the probe to total DNA from group of bacteria ecologically similar to P. tolaasii including WLORs, fluorescent Pseudomonads isolated from oyster mushroom, P. agarici, P. gingeri, and some of other species of Psedomonas showed that some of the tested bacteria do not have any hybridized band and others have bands sowing RFLP. The cloned DNA fragment or its nucleotide sequence will be useful in detection and identification of the P. tolaasii.

• 환경생물
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• 제22권1호
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• pp.213-219
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• 2004

동굴 내 정점별 세균 군집 구조를 분석하기 위하여 PCf amplified 16S rDNA denaturing gradient gel electrophoresis(DGGE)를 적용하였다. DGGE는 동일한 분자량을 갖는 dsDNA band라고 할지라도, 각각의 염기서열 차이에 따라 전기영동 상에서 고유한 band양상을 나타낼 수 있다. eubacteria의 16S rDNA V3region을 증폭하기 위해 GC341F와 PRUN518r을 primer로 사용하여 지하수내에 미생물 군집의 다양성과 유사성을 분석하였다. DGGE band 양상을 통해 동굴내의 세균 군집 구조는 외부 환경에 비해 상대적으로 종다양성이 낮으며 동굴내 에서 특이적으로 서식하는 종이 있음을 확인하였다. 또한 유기 영양물질의 공급이 제한되어 있는 동굴에서 구아노가 주요 유기 영양물질의 공급원으로서 큰 영향을 미치고 있는 것으로 파악되었다. DGGE 상의 일부 band의 염기서열분석 결과 Pseudomonas sp. NZ060과 Pseudomonas pseudoalcaligenes, uncultured Variovorax sp., soil bacterium NS7로 동정되었다.

• Journal of Plant Biotechnology
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• 제42권3호
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• pp.154-160
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• 2015

The plant breeding technology was developed with genetic engineering. Many researchers and breeders have turned from traditional breeding to molecular breeding. Genetically modified organisms (GMO) were developed via molecular breeding technology. Currently, molecular breeding technologies facilitate efficient plant breeding without introducing foreign genes, in virtue by of gene editing technology. Gene targeting (GT) via homologous recombination (HR) is one of the best gene editing methods available to modify specific DNA sequences in genomes. GT utilizes DNA repair pathways. Thus, DNA repair systems are controlled to enhance HR processing. Engineered sequence specific endonucleases were applied to improve GT efficiency. Engineered sequence specific endonucleases like the zinc finger nuclease (ZFN), TAL effector nuclease (TALEN), and CRISPR-Cas9 create DNA double-strand breaks (DSB) that can stimulate HR at a target site. RecQl4, Exo1 and Rad51 are effectors that enhance DSB repair via the HR pathway. This review focuses on recent developments in engineered sequence specific endonucleases and ways to improve the efficiency of GT via HR effectors in plants.

• Journal of Plant Biotechnology
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• 제44권3호
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• pp.235-242
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• 2017

Cudrania tricuspidata Bureau is a widely-used, medicinal, perennial and woody plant. Obtaining information about the genetic diversity of plant populations is highly important with regard toconservation and germplasm utilization. Although C. tricuspidata is an important medicinal plant species registered in South Korea, no molecular markers are currently available to distinguish Korean-specific ecotypes from other ecotypes from different countries. In this study, we developed single nucleotide polymorphism (SNP) markers derived from the chloroplast and nuclear genomic sequences, which serve to to identify distinct Korean-specific ecotypes of C. tricuspidata via amplification refractory mutation system (ARMS)-PCR and high resolution melting (HRM) curve analyses. We performed molecular authentication of twelve C. tricuspidata ecotypes from different regions using DNA sequences in the maturaseK (MatK) chloroplast intergenic region and nuclear ribosomal DNA internal transcribed spacer (ITS) regions. The SNP markers developed in this study are useful for rapidly identifying specific C. tricuspidata ecotypes from different regions.

• 한국식물학회:학술대회논문집
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• 한국식물학회 1987년도 식물생명공학 심포지움 논문집 Proceedings of Symposia on Plant Biotechnology
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• pp.149-155
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• 1987

Growth and development of a higher plant, or any living organism for that matter, could be defined as an orderly expression of the genome in time and space in close interaction with the environment. During differentiation and development of a tissue or organ a group of genes must be selectively turned on or turned off mainly by trans-acting regulators. In this general concept of regulation of regulation of gene expression, a DNA molecule is recognized at a specific nucleotide sequence by DNA-binding factors. Molecular biology of the regulatory factors such as hormones, and their receptors, target DNA sequences and DNA-binding proteins are well advanced. What is not clearly understood is the molecular basis of the interactions between DNA and binding factors, expecially of the usages of the dyad symmetry of the target DNA sequences and the dimeric nature of the DNA-binding proteins. A unique 3-dimensional structure of DNA has been proposed that may play an important role in the orderly expression of the gene. A foldback intercoil (FBI) DNA configuration which was originally found by electron microscopy among mtDNA molecules from pearl millet has some unique features. The FBI configuration of DNA is believed to be formed when a flexible double helix folds back and interwines in the widened major grooves resulting in a four stranded, intercoil DNA whose thickness is the same as that of double stranded DNA. More recently, the FBI structure of DNA has been also induced in vitro by a novel enzyme which was purified from pearl millet mitochondria. It has been proposed that the FBI DNA could be utillized in intramolecular recombination which leads to inversion or deletion, and in intermolecular recombination which can lead to either site-specific recombination, genetic recombination via single strand invasion, or cross strand recombination. The structure and function of DNA in 3-dimensional aspect is emphasized for better understanding orderly expression of genes during growth and development.

• Journal of Microbiology and Biotechnology
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• 제15권6호
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• pp.1377-1383
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• 2005

In a cDNA microarray experiment using Cy3 and Cy5 as labeling agents, particularly for the direct design, cDNAs from some genes incorporate one dye more efficiently than the other, which is referred to as the gene-specific dye bias. Dye-swaps, in which two dyes are switched on replicate arrays, are commonly used to control the gene-specific dye bias. We developed a simple procedure to extract the gene-specific dye bias information from a partial dye swap experiment. We detected gene-specific dye bias by identifying outliers in an X-Y plane, where the X axis represents the average log-ratio from two sets of dye swap pairs and the Y axis exhibits the average log ratio of four forward labeled arrays. We used this information for detecting differentially expressed genes, of which the additionally detected genes were validated by real-time RT-PCR.

• Mycobiology
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• 제31권4호
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• pp.196-199
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• 2003

A very reliable and specific method for the identification of fungi in ectotrophic mycorrhizal symbiosis was developed using a specific PCR assay based on the amplification of the ITS1 region. To obtain specific data, an ITS-diagnostic assay was carried out that reveals genera and species specific sequences. Here, an application of one method is presented, which covers the identification of pure mycelia, basidiocarps as well as mixed samples such as ectomycorrhizal roots that were mingled with remains of the host plant. For this purpose a protocol was established that allowed the extraction of DNA from single mycorrhizal roots. In order to perform a specific ITS analysis we generated a new ITS-primer(ITS8) by a multiple alignment of five different genera and species of mycorrhizal fungi. The utilization of ITS1 and ITS8 resulted in specific PCR amplicons, which were characterized by sequencing without purification steps, even when the template DNA was associated with roots.

• 대한수의학회지
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• 제45권3호
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• pp.351-357
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• 2005

Here I describe a multiplex allele specific PCR-based approach for the rapid detection between Hanwoo and Holstein meat associated with Melanocortin 1 receptor (MC1R) gene. Specific and universal oligonucleotide primers were used in combination to detect the presence of a single nucleotide polymorphism within the bovine MC1R DNA sequence. The presence of the bovine MC1R gene is indicated by the production of a single control PCR product, whilst positive samples generate an alternative smaller specific product over the same region. The mutations in MC1R104 codon revealed depending on the presence or absence of an indicative fragment amplified from the wild-type allele of this codon. As little as 0.39 ng and 1.56 ng of genomic DNA of Hanwoo and Holstein could be detected by MAS-PCR assay, respectively. This technique, which is widely used in human genetic screening, provides a reliable and sensitive result that has not been documented for the identification of bovine coat color. The MAS-PCR assay approach was proven to be useful in complementing routine beef DNA analysis for differentiation of these MC1R variants and it would facilitate the screening of deceiving sales of Holstein meat in the butcher shop.