• Korean journal of applied entomology
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• v.36 no.1
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• pp.30-36
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• 1997

Except for a cosmopolitan and major pest of apples, Tetranychus urticae Koch, Tetranychus mites in Korea such as T. viennensis Zaher, T. kanzawai Kishida, and T. truncams Ehara have been considered as quarantine pests by Canada and United States. Even though these mites are not feeders on apples, they are suspected to attach accidentally on apple h i t s in autumn as females enter the diapause. The characters used to identify Tetranychus mites have been confined to the shape of aedeagus in adult male. To develope a fast and accurate alternative identification protocol applied to hibernating female mites on apples, their mitochondrial DNA (mtDNA) were examined to find out any polymorphisms to discriminate each species from the other ones. Three pairs of primers for polymerase chain reaction (PCR) were used to amplify cytochrome oxidase subunit I (CO-I) coding region in mitochondrial DNA5 of four species of Tetranycus mites. The longest amplified product was estimated its size as about 680 bp. Digestion with restriction enzymes, AluI, Ddel, and Sau3A, showed length polymorphisms, which will he useful as diagnostic markers to identify Tetranychus mites. Schematic restriction maps in amplified region were shown for each species.

• Korean Journal of Soil Science and Fertilizer
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• v.33 no.4
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• pp.266-274
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• 2000

Understanding of microbial community structure in soil-root system is necessary to use beneficial soil and rhizosphere microbes for improvement of crop production and biocontrol. The knowledge of behavior and function of microbes in soil-root system plays a key role for the application of beneficial inocula. Because the majority of the intact bacteria in soil are unable to grow on nutrient media, both culturable and nonculturable bacteria have to be studied together. In our study, culture-independent survey of bacterial community in the soil-root system of red pepper fields was conducted by the sequence analysis of three universal clone libraries of genes which code for small-subunit rRNA (rDNA). Universal small subunit rRNA primers were used to amplify DNA extracted from each sample and PCR products were cloned into pGEM-T. Out of 27 clones sequenced, 25 clones were from domain bacteria. Two of the rDNA sequences were derived from eukaryotic organelles. Within the domain bacteria, several kingdoms were represented : the Proteobacteria (16 clones). Cytophyga-Flexibacter-Bacteroides group (2 clones). the high G+C content gram-positive group(1 clone) and 4 unknown clones.

• Research in Plant Disease
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• v.15 no.3
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• pp.165-169
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• 2009

Twelve Cucumber mosaic virus (CMV) isolates were isolated from genetically modified (GM) and non-GM Capsicum annuum in two GM fields, Namyangju and Anseong, and their properties were investigated in this study. Coat protein (CP) gene of the CMV isolates were synthesized by RT-PCR using genus-specific primers which designed to amplify a DNA fragment of 950 bp. Purified cDNA fragments were cloned into the pGEMT easy vector for sequence determination. Nucleotide sequences (internal 657 bp) of CMV isolates were compared with Fny-CMV CP sequences and there were no significant collection site specific sequence similarities found. When predicted amino acid sequences (219 amino acids) were compared with Fny-CMV CP amino acids sequences, there were 96.8% to 97.3% similarities found from Namyangju collections and 95.9% to 96.8% similarities from Anseong collections. The phylogenetic analysis with nucleotide sequences showed definite differences in CMVs which have been isolated from the two regions.

• Clinical and Experimental Reproductive Medicine
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• v.24 no.2
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• pp.267-271
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• 1997

Apoptosis, programmed cell death, is posulated to occur in granulosa cells in ovarian follicular atresia. bcl-2 gene serves as protector from apoptosis and, thus, is associated with increased cell survival. TRPM-2 gene expression has been implicated as a trigger of apoptosis in rat prostate, uterus and mammary gland. Our objective was to determine if bcl-2 and TRPM-2 are expressed in luteinized human GC and, therefore, have regulatory functions for apoptosis in GC. Human GC were obtained via oocyte retrival from the infertile patients stimulated with exogeneous gonadotropins while undergoing IVF. GC were isolated from follicular fluid using Percoll gradient centrifugation. The GC were further purified with anti-CD45 magnetic beads to remove contaminating WBC's. RT-PCR were performed to analyze the mRNA expression of bcl-2 and TRPM-2 in the GC. The PCR primers were designed to amplify a 195 bp fragment of bcl-2 and a 174 bp fragment of TRPM-2. The PCR products were electrophoresed on 4% agarose gel. Three separate experiments indicated that both bcl-2 and TRPM-2 are concurrently expressed in human GC. We cultured granulosa cells with FSH (1 ng/ml) for 1 day to investigate the relative changes of TRPM-2 mRNA level with RNAse protection assay. When we cultured GC with serum free medium for 1 day TRPM-2 mRNA level increased with 1.3 fold, however it was decreased 0.64 fold with FSH. Therefore we conclude that bcl-2 and TRPM-2 are concurrently expressed and that the interaction of their products may be involved in GC apoptosis. And TRPM-2 may be regulated with FSH.

• The Korea Journal of Herbology
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• v.24 no.4
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• pp.115-120
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• 2009

Objectives : Due to the morphological similarity and frequent occurrence of intermediate forms as well as morphological variations of aerial part, the correct identification between Rhei Radix et Rhizoma and Rhei Undulatai Rhizoma is very difficult. To develop a reliable method for correct identification and improving the quality standards of Rhei Radix et Rhizoma and Rhei Undulatai Rhizoma, we analyzed RAPD and developed SCAR marker. Methods : To amplify target DNA at the genomic level, 32 Operon 10-mer random primers were applied with four Rheum species, R. officinale, R. palmatum, R. tanguticum and R. undulatum. The nucleotide sequences were determined and species-specific primers were prepared depending on the species-specific RAPD amplicons after subcloned into the pGEM-Teasy vector. To develop the SCAR markers, species-specific PCR amplification and multiplex-PCR were carried out using the single species-specific primer pairs and combinations of them, respectively. Results : We used RAPD analysis of four Rheum plant species to obtain several species-specific RAPD amplicons. From nucleotide sequences of these RAPD amplicons, we developed two SCAR markers that amplified 314 bp and 390 bp DNA fragments in only R. undulatum but not in R. officinale, R. palmatum, R. tanguticum and R. undulatum, for distinguishing Rhei Undulatai Rhizoma and Rhei Radix et Rhizoma. Furthermore, we established SCAR markers for the simultaneous discrimination of the three species within a single reaction by using multiplex-PCR. Conclusions : These genetic markers can be used for the efficient discrimination of plants species and commercial herbal medicines between Rhei Undulatai Rhizoma and Rhei Radix et Rhizoma, to ultimately prevent indiscriminate distribution and prescription of these herbal medicines.

• International Journal of Industrial Entomology and Biomaterials
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• v.18 no.1
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• pp.18-27
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• 2009

B. thuringiensis strain LY-99 belonging to subsp. alesti (H3a3c), was isolated from Chinese tobacco warehouse and showed significantly high toxicity to Plutella xylostella. For the identification of the cry1-type genes from B. thuringiensis LY-99, an extended multiplex PCRrestriction fragment length polymorphism (PCRRFLP) method was established by using two pairs of universal primers based on the conserved regions of the cry1-type genes to amplify around 2.4 kb cry1-type gene fragments. Then the DNA fragment was cloned into pGEM-T Easy vector and digested with EcoRI and EcoRV enzymes. Through this method, a known cry1-type gene was successfully identified from the reference strain, B. thuringiensis subsp. alesti. In addition, the RFLP patterns revealed that B. thuringiensis LY-99 included a novel cry1A-type gene in addition to cry1Aa, cry1Ac, cry1Be and cry1Ea genes. The novel cry1A-type gene was designated cry1Ah2 (Genbank accession No DQ269474). An inverse PCR method was used to amplify the flank regions of cry1Ah2 gene. Finally, 3143 bp HindIII fragment from B. thuringiensis LY-99 plasmid DNA including 5' region and partial ORF was amplified, and sequence analysis revealed that cry1Ah2 gene from LY-99 showed 89.31% of maximum sequence similarity with cry1Ac1 crystal protein gene. In addition, the deduced amino acid sequence of Cry1Ah2 protein shared 87.80% of maximum identity with that of Cry1Ac2. This protein therefore belongs to a new class of B. thuringiensis crystal proteins.

• Journal of Life Science
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• v.13 no.5
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• pp.723-731
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• 2003

For the investigation of conserved regions in lipase genes, 132 and 24 sequences were obtained from LED (Lipase Engineering Database) and COG (Clusters of Orthologous Groups of proteins), respectively. There was high diversity in lipase genes and peculiar amino acid sequences were conserved for each homologous family of LED. Similar conserved amino acid sequences were detected from COG0657 and Moraxella lipase 1 homologous group of LED. Although many studies have attempted to detect new lipase genes in procaryotes, they have been limited culturable bacteria. The importance of metagenome, including DNA from non-culturable bacteria, is known. Due to the high diversity, we assumed it might be possible to detect new lipase gene from metagenome. Due to the high diversity of nucleotide sequences in lipase genes, 10 primer sets were designed. Designed primer sets were inspected in BLAST of NCBI and they could amplify a part of the lipase gene from 222 to 713 bp. They can amplify 16.7%∼60.0% of each lipase homologous group which was 3.6 fold higher than each sets. They might offer a high probability of detecting new lipase genes, owing to high efficiency and the diversity of lipase genes.

• Journal of Food Hygiene and Safety
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• v.27 no.2
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• pp.182-187
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• 2012

In this study, the experimental method has been investigated using molecular biological way to identify raw materials from seasoned red-pepper sauce which is one of the most popular spices in Korea. 6 kinds of seasoned red-pepper sauces were chosen as a sample containing chilli pepper, garlic, onion as a major ingredient and species specific primers were used for the identification of the raw material of processed food. Selected samples were pre-treated to remove salt (samples were washed with distilled water 3~4 times for desalting), after that, to amplify the extracted genes, whole genome amplification (WGA) kit was performed. Afterwards, PCR products were confirmed through the electrophoresis. As a result, 102, 180, 280 bp of specific PCR products were confirmed for each major ingredients such as chilli pepper, garlic, onion. From this study, the gene extraction method was validated for the identification of ingredients from the spices and it would be applied to distinction of low quality chilli pepper powder including seasoned red-pepper sauce illegally.

• Asian-Australasian Journal of Animal Sciences
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• v.12 no.5
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• pp.689-694
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• 1999

This experiment was carried out to isolate the partial bovine (Bos Taurus coreanae) myogenic factor encoding gene, MyoD, using the rat myogenic factor (MyoD) gene sequence and to compare the gene sequence between another myogenic factor (Myf 5) and MyoD gene of the bovine. To make the probe and isolate the MyoD gene, PCR was performed to amplify rat and bovine MyoD gene including exon I, II and intron I. The homology between mouse and bovine MyoD is high; bovine MyoD gene shows 17 different gene sequence region compared to rat MyoD. Among those, two regions have significant differences; one is the exon I part between 2834 and 2850 bp, the other is intron part between 3274 and 3303 bp of the mouse. At this region homology was 40% in the former and 50% in the latter. Homology between bovine MyoD and Myf5 was 83% in the exon 1. Especially exon I in the Myf5 602-617 bp and 651-683 bp have significant differences. These results suggest that MyoD gene have a similar gene structure in mouse and bovine and MyoD and Myf5 of the bovine, at least in part, have a similar expression and activity.

• Journal of Forest and Environmental Science
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• v.25 no.2
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• pp.93-100
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• 2009

In Malaysia, Labisia pumila Benth & Hook f, popularly known as 'Kacip Fatimah' has been used traditionally to treat various elements of the woman's health in Malay community. The objective of this study was to develop randomly amplified polymorphic DNA (RAPD) based DNA markers for the identification of L. pumila and to distinguish its three varieties from each other. Total DNA from nine accessions of L. pumila was extracted by CTAB method and polymerase chain reactions (PCR) were carried out to amplify the segments of DNA using different primers to develop DNA barcode using RAPD technique. To find out variety-specific DNA marker/s, twenty different 10-mer primer sequences with annealing temperature from 36-$40^{\circ}C$ were evaluated in triplicate. Out of 20 random primers, two primers (OPA-1 and OPA-2/A10) were selected which produced reliable RAPD band patterns. To have DNA based handle, two RAPD amplification products were cloned and sequenced to determine the identity of the DNA. RAPD analysis using two random primers generated 72 discrete bands ranging in size 200 bp-3,000 bp. Fifty nine of these were polymorphic loci (82%) and thirteen were non-polymorphic loci (18%). A total of 32 bands polymorphic loci (72%) were amplified with primer OPA-1 and analyzed by cluster analysis and UPGMA (Unweighted Pair Group Method with Arithmetic) to present a dendogram depicting the degree of genetic relationship among nine accessions of L. pumila. Our results shows the reasonable genetic diversity among the L. pumila varieties and within varieties; and two RAPD marker sequences obtained could be used to identify L. pumila at species level.