• Restorative Dentistry and Endodontics
• /
• v.33 no.3
• /
• pp.224-234
• /
• 2008

The purpose of this study was to compare the normal and two times of application time of six self-etching primers applied to enamel using microshear bond strength (uSBS) test and the finding of scanning electronic microscope (SEM). Crown of sixty human molars were bisected mesiodistally and buccal and lingual enamel of crowns were partially exposed and polished with 600 grit SiC papers. They were divided into one of two equal groups subdivided into one of six equal groups (n = 10) by self-etching primer adhesives. After the same manufacture's adhesive resin and composites were bonded on the enamel surface of each group, the bonded specimens were subjected to uSBS testing and also observed under SEM. In conclusion, generally two times of primer application time increased the enamel uSBS, especially with the statistical increase of bond strength in adhesives involving high-pH primers.

• FLOWER RESEARCH JOURNAL
• /
• v.18 no.3
• /
• pp.201-206
• /
• 2010

The genetic relationship among 48 Cymbidium cultivars was analyzed using randomly amplified polymorphic DNA (RAPD) with eighty 10 mers random primers (Operon Technologies) and twelve 20 mers random primers. Forty eight Cymbidium cultivars included 34 oriental Cymbidium, 7 hybrids, and 7 western Cymbidium. 407 (9.9 per primer) and 56 polymorphic bands (9.5 per primer) were generated by polymerase chain reaction with selected thirty 10 mers primers, and nine 20 mers primers, respectively. The polymorphic fragments ranged from 0.4 to 1.5 kb in size. The dendrogram was constructed by using the UPGMA clustering algorithm based on genetic similarity. Forty eight Cymbidium cultivars were classified into four major groups at similarity coefficient value of 0.638.

• The Plant Pathology Journal
• /
• v.34 no.6
• /
• pp.490-498
• /
• 2018

Panicle blight and seed rot disease caused mainly by Burkholderia glumae and Burkholderia gladioli is threatening rice cultivation worldwide. The bacteria have been reported as seed-borne pathogens from rice. Accurate detection of both pathogens on the seeds is very important for limiting the disease dissemination. Novel primer pairs targeting specific molecular markers were developed for the robust detection of B. glumae and B. gladioli. The designed primers were specific in detecting the target species with no apparent cross-reactions with other related Burkholderia species at the expected product size. Both primer pairs displayed a high degree of sensitivity for detection of B. glumae and B. gladioli separately in monoplex PCR or simultaneously in duplex PCR from both extracted gDNA and directly preheated bacterial cell suspensions. Limit of detection was as low as 0.1 ng of gDNA of both species and $3.86{\times}10^2cells$ for B. glumae and $5.85{\times}10^2cells$ for B. gladioli. On inoculated rice seeds, the designed primers could separately or simultaneously detect B. glumae and B. gladioli with a detection limit as low as $1.86{\times}10^3cells$ per rice seed for B. glumae and $1.04{\times}10^4cells$ per rice seed of B. gladioli. The novel primers maybe valuable as a more sensitive, specific, and robust tool for the efficient simultaneous detection of B. glumae and B. gladioli on rice seeds, which is important in combating rice panicle blight and seed rot by early detection and confirmation of the dissemination of pathogen-free rice seeds.

• The Plant Pathology Journal
• /
• v.35 no.2
• /
• pp.172-177
• /
• 2019

A duplex PCR method was developed for simultaneous detection and identification of tobacco root rot pathogens Phytophthora nicotianae and Thielaviopsis basicola. The specific primers for P. nicotianae were developed based on its internal transcribed spacer (ITS) regions of ribosomal gene, ras gene and hgd gene, while the specific primers for T. basicola were designed based on its ITS regions and ${\beta}$-tubulin gene. The specificity of the primers was determined using isolates of P. nicotianae, T. basicola and control samples. The results showed that the target pathogens could be detected from diseased tobacco plants by a combination of the specific primers. The sensitivity limitation was $100fg/{\mu}l$ of pure genomic DNA of the pathogens. This new assay can be applied to screen out target pathogens rapidly and reliably in one PCR and will be an important tool for the identification and precise early prediction of these two destructive diseases of tobacco.

• Research in Plant Disease
• /
• v.29 no.1
• /
• pp.94-99
• /
• 2023

Blueberry red ringspot virus (BRRSV) and blueberry scorch virus (BlScV) are included in the quarantine virus list managed by the Korean Animal and Plant Quarantine Agency. A multiplex polymerase chain reaction (PCR) assay with an internal control was developed for the simultaneous detection of both viruses. The specific primers used here were designed based on the highly conserved regions of the genomic sequences of each virus, obtained from the National Center for Biotechnology Information nucleotide databases. The primers were designed to amplify a partial sequence within coat protein (CP) for detecting BRRSV and a partial sequence within the CP-16 kDa for detecting BlScV. 18S ribosomal RNA (rRNA) was used as internal control, and the primer set used in a previous study was modified in this study for detecting 18S rRNA. Each conventional PCR using the BRRSV, BlScV, and 18S rRNA primers exhibited a sensitivity of approximately 1 fg plasmid DNA. The multiplex PCR assay using the BRRSV, BlScV, and 18S rRNA primers was effective in simultaneously detecting the two viruses and 18S rRNA with a sensitivity of 1 fg plasmid DNA, similar to that of conventional PCR assays. The multiplex PCR assay developed in this study was performed using 14 blueberry cultivars grown in South Korea. BRRSV and BlScV were not detected, but 18S rRNA was all detected in all the plants tested. Therefore, our optimized multiplex PCR assay could simultaneously detect the two viruses and 18S rRNA in field samples collected from South Korea in a time-efficient manner. This approach could be valuable in crop protection and plant quarantine management.

• Mycobiology
• /
• v.31 no.2
• /
• pp.68-73
• /
• 2003

Roots of Glycine max and Miscanthus sinensis and soil samples were collected from various field sites at Goesan, Chungbuk in Korea. Microscopic observations of the roots indicated high colonization rates of both arbuscular mycorrhizal fungi(AMF) and other fungi. The partial small subunit of ribosomal DNA genes were amplified with the genomic DNA extracted from their roots by nested polymerase chain reaction(PCR) with universal primer NS1 and fungal specific primers AML Restriction fragment length polymorphism(RFLP) was analyzed using the combinations of three restriction enzymes, HinfI, AluI and AsuC21. Nucleotides sequence analysis revealed that ten sequences from Miscanthus sinensis and one sequence from Glycine max were close to those of arbuscular mycorrhizal fungi. Also, 33% of total clones amplified with NS31-AM1 primers from M. sinensis and 97% from G. max were close to Fusarium oxysporum or other pathogenic fungi, and they were successfully distinguished from AME Results suggested that these techniques could help to distinguish arbuscular mycorrhizal fungi from root pathogenic fungi in the plant roots. Especially, DNA amplified by these primers showed distinct polymorphisms between AMF and plant pathogenic species of Fusarium when digested with AsuC21.

• Asian Pacific Journal of Cancer Prevention
• /
• v.17 no.10
• /
• pp.4741-4745
• /
• 2016

Background and Objective: Any role of human papillomavirus (HPV) in the development of breast cancer is conjectural. The aim of this study was to investigate possible links between HPV and breast cancer in women, Sanandaj, Iran. Methods: In this case-control study, 70 formalin fixed and paraffin embedded blocks of breast malignant tumors as a case group and 70 blocks of lesions without malignancy were selected as controls. Sections about $10{\mu}m$ thick were prepared. After removing the paraffin, DNA was extracted. Samples were tested by PCR using general and high-risk specific HPV primers. Results: All 70 malignant breast tumors (cases) were invasive ductal carcinomas, and of the 70 controls, 17 (24.3%) were fibrocystic tumors and 53 (75.7%) fibroadenomas. The age range of women in the case group was 25-72 years old and in the control group It was13-66 years. Using HPV general primers two samples were positive in the case group, confirmed to be HPV-18 using high-risk specific primers. Conclusion: No statistically significant association was found between breast cancer and HPV. It is necessary to confirm this result by further investigations in other populations.

• BMB Reports
• /
• v.36 no.5
• /
• pp.427-432
• /
• 2003

Jute is the principal coarse fiber for commercial production and use in Bangladesh. Therefore, the development of a high-yielding and environmental-stress tolerant jute variety would be beneficial for the agro economy of Bangladesh. Two molecular fingerprinting techniques, random-amplified polymorphic DNA (RAPD) and amplified-fragment length polymorphism (AFLP) were applied on six jute samples. Two of them were cold-sensitive varieties and the remaining four were cold-tolerant accessions. RAPD and AFLP fingerprints were employed to generate polymorphism between the cold-sensitive varieties and cold-tolerant accessions because of their simplicity, and also because there is no available sequence information on jute. RAPD data were obtained by using 30 arbitrary oligonucleotide primers. Five primers were found to give polymorphism between the varieties that were tested. AFLP fingerprints were generated using 25 combinations of selective-amplification primers. Eight primer combinations gave the best results with 93 polymorphic fragments, and they were able to discriminate the two cold-sensitive and four cold-tolerant jute populations. A cluster analysis, based on the RAPD and AFLP fingerprint data, showed the population-specific grouping of individuals. This information could be useful later in marker-aided selection between the cold-sensitive varieties and cold-tolerant jute accessions.

• The Plant Pathology Journal
• /
• v.18 no.1
• /
• pp.12-17
• /
• 2002

For the molecular detection of Sweet potaio feathery mottle virus (SPFMV) from diseased sweet potato plants, reverse transcription and polymerase chain reaction (RT-PCR) was performed with the use of a set of virus-specific primers to amplify an 816 bp product. The viral coat protein gene was selected for the design of the primers. No PCR product was amplified when Turnip mosaic virus, Potato vims Y or Cucumber mosaic virus were used as template in RT-PCR with the SPFMV-specific primers. The lowest concentration of template viral RNA required for detection was 10 fg. The vim was rapidly detected from total nucleic acids of leaves and roots from the virus-infected sweet potato plants as well as from the purified viral RNA by the RT-PCR. Twenty-four sweet potato samples were selected and analyzed by RT-PCR and restriction fragment length polymorphism (RFLP). RFLP analysis of the PCR products showed three restriction patterns, which resulted in some point mutations suggesting the existence of quasi-species for the vims in the infected sweet potato plants.

• 한국신재생에너지학회:학술대회논문집
• /
• 2005.11a
• /
• pp.631-633
• /
• 2005

For biohydrogen production, hydrogenase is a key enzyme. In the present work we performed search of [NiFe]-hydrogenases from hydrogen producing microorganisms using degenerate polymerase chain reaction (PCR) strategy. Degenerate primers were designed from the conserved region of [NiFe]-hydrogenase group I especially on structural genes encoding for catalytic subunit of [NiFe]-hydrogenase from bacteria producing hydrogen. Most of [NiFe]-hydrogenase (group I) are expressed via complex mechanism with aid of auxiliary protein and localized through twin-arginine translocation pathway. [NiFe]-hydrogenase is composed of large and small subunits for catalytic activity. It is known that only small subunit has signal peptide for periplasmic localization and large & small subunitscome together before localization. During this process, large subunit is treated by endopeptidase for maturation. Based on these information we used signal peptide sequence and C-terminal of large subunit by recognized by endopeptidase as templates for degenerate primers. About 2,900 bp of PCR products were successfully amplified using the designed degenerate primers from genomic DNAs of several microorganisms. The amplified PCR products were inserted into T-vector and then sequenced to confirm.