• The Plant Pathology Journal
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• v.37 no.5
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• pp.476-488
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• 2021

Xanthomonas campestris pv. campestris (Xcc) is the causal agent of black rot for cruciferous vegetables worldwide, especially for the cole crops such as cabbage and cauliflower. Due to the lack of resistant cabbage cultivars, black rot has brought about considerable yield losses in recent years in China. Understanding of the pathogen features is a key step for disease prevention, however, the pathogen diversity, population structure, and virulence are largely unknown. In this study, we studied 50 Xcc strains including 39 Xcc isolates collected from cabbage in 20 regions across China, using multilocus sequence genotyping (MLST), repetitive DNA sequence-based PCR (rep-PCR), and pathogenicity tests. For MLST analysis, a total of 12 allelic profiles (AP) were generated, among which the largest AP was AP1 containing 32 strains. Further cluster analysis of rep-PCR divided all strains into 14 DNA groups, with the largest group DNA I comprising of 34 strains, most of which also belonged to AP1. Inoculation tests showed that the representative Xcc strains collected from diverse regions performed differential virulence against three brassica hosts compared with races 1 and 4. Interestingly, these results indicated that AP1/DNA I was not only the main pathotype in China, but also a novel group that differed from the previously reported type races in both genotype and virulence. To our knowledge, this is the first extensive genetic diversity survey for Xcc strains in China, which provides evidence for cabbage resistance breeding and opens the gate for further cabbage-Xcc interaction studies.

• Reproductive and Developmental Biology
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• v.34 no.3
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• pp.181-184
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• 2010

Osmolarity of culture media is one of the most important factors affecting in vitro development. This study was conducted to investigate the DNA methylation status of Pre-1 and satellite sequence in pig nuclear transfer (pNT) embryos produced under different osmolarity culture conditions. Control group of pNT embryos was cultured in PZM-3 for six days. Other two treatment groups of pNT embryos were cultured in modified PZM-3 with 138 mM NaG or 0.05M sucrose (mPZM-3, 320 mOsmol) for two days, and then cultured in PZM-3 (270 mOsmol) for four days. Previous our studies have reported that pNT embryos cultured in both hypertonic media showed significantly higher blastocyst formation rate than that of control. The DNA methylation status of the satellite sequences in blastocyst was characterized using bisulfite-sequencing technology. The satellite region had a similar methylation pattern of in vivo blastocyst among two culture groups excepting the control group. Each level of methylation is that the satellite DNA moderately methylated (43.10% of PZM-3; 56.12% of NaCl; 55.06% of sucrose; 60.00% of in vivo embryos). As a result of the sequence of PRE-1, CpG methylation pattern was similar to three groups, including in vivo group. In case of the satellite DNA region, the osmolarity conditions were affected CpG DNA methylation status while PRE-1 sequence was not affected CpG DNA methylation in pNT blastocyst stage. These results indicate that the modification of osmolarity in a culture media may influence to spatially change of DNA methylation of repetitive sequence for pNT embryo development.

• Korean Journal of Agricultural Science
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• v.38 no.2
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• pp.269-275
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• 2011

The objective of this study was to develop a rapid and reliable PCR method for sexing of morula or blastocyst stage bovine embryo. BOV97M and bovine 1.715 satellite DNA sequences were selected for amplification of male and bovine specific DNA, respectively. But the unbalanced number of copies of these two repetitive sequences required some modification of PCR method. Karyotyping of blastomeres were carried for the confirmation of sex determination in bovine embryos. The coincidence rate of sex between biopsied-single blastomere and matched blastocyst was 80.0%. When in vivo- and in vitro- derived embryos were compared, 61.8% and 56.7% were male in in vitro- and in vivo-derived embryos, respectively. In vivo-derived embryos showed better hatching rate than in vitro-derived embryos following biopsy of blastomeres. In conclusion, rapid and effective PCR could be applied to sexing of bovine preimplantation embryos using single blastomere. The sensitivity of this assay may eliminate the need for biopsy of more than one nucleated blastomere and reduce trauma to the embryos derived from biopsy procedure.

• Journal of Microbiology
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• v.42 no.2
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• pp.87-93
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• 2004

Twelve mecoprop-degrading bacteria were isolated from soil samples, and their genetic and phenotypic characteristics were investigated. Analysis of 16S rDNA sequences indicated that the isolates were related to members of the genus Sphingomonas. Ten different chromosomal DNA patterns were obtained by polymerase-chain-reaction (PCR) amplification of repetitive extragenic palindromic (REP) sequences from the 12 isolates. The isolates were found to be able to utilize the chiral herbicide meco-prop as a sole source of carbon and energy. While seven of the isolates were able to degrade both (R)-and (S)-mecoprop, four isolates exhibited enantioselective degradation of the (S)-type and one isolate could degrade only the (R)-enantiomer. All of the isolates were observed to possess plasmid DNAs. When certain plasmids were removed from isolates MPll, MP15, and MP23, those strains could no longer degrade mecoprop. This compelling result suggests that plasmid DNAs, in this case, conferred the ability to degrade the herbicide. The isolates MP13, MP15, and MP24 were identified as the same strain; however, they exhibited different plasmid profiles. This indicates that these isolates acquired dif-ferent mecoprop-degradative plasmids in different soils through natural gene transfer.

• The Korean Journal of Mycology
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• v.27 no.2 s.89
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• pp.132-137
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• 1999

This study was carried out to develop the molecular marker for the detection of Pseudomonas tolaasii, a causative agent of bacterial brown blotch disease of oyster mushroom (Pleurotus ostreatus). When several primers designed from repetitive sequences and pectin lyase genes of bacteria were used to produce DNA polymorphism from different Pseudomonas spp. isolated from edible mushrooms, PEU1 primer derived from pectin lyase gene produced polymorphic bands differentiating P. tolaasii strains from other Pseudomonas species. Two bands, 1.0kb and 0.4kb, found commonly in 6 isolates of P. tolaasii were cloned into pGEM-T vector which were designated as pPTOP1 and pPTOP2, respectively, to use as probe. The 0.4 kb insert of pPTOP2 hybridized to only 6 isolates of P. tolaasii, but did not to the other Pseudomonas species. As few as $1.5{\times}10^3$ colony forming unit (cfu) of P. tolaasii could be detected by dot blot hybridization with the cloned 0.4kb DNA in pPTOP2.

• Journal of Food Hygiene and Safety
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• v.18 no.4
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• pp.229-236
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• 2003

The five different foodborne pathogenic bacterial genera of Escherichi, Salmonella, Shigella, Vibrio and Listeria are important sources of foodpoison. However, the method was not developed to simultaneously differentiate these five bacteria at molecular level. The optimized concentrations of the four major PCR cocktail components of $MgCl_2$, dNTPs, primers and template DNA were determined when ERIC (enterobacterial repetitive intergenic consensus)-PCR reactions were carried out to differentiate the five differnet foodborne pathogenic bacteria. The optimized concentration of $MgCl_2$ was determined to be 2 mM in order to obtain a consistent fingerprinitng pattern. The similar fingerprinting pattern was obtained when ERIC primers and dNTPs were added up to the concentrations of 2 ${\mu}M$ and 200 ${\mu}M$, respectively. As for template DNA, the numbers of PCR fragments were not affected, but their intensities were increased as the concentrations of the DNA were increased.

• Clinical and Experimental Reproductive Medicine
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• v.27 no.4
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• pp.405-415
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• 2000

Objective: The present study was undertaken to synthesize a human Y chromosome specific probe and to confirm the usefulness of the probe for fluorescence in situ hybridization (FISH) in various types of human cells. Methods: An approximately 400 bp DNA fragment of the DYZ1 sequences was synthesized by PCR using digoxigenin labeled dUTP (dig-PCR). The fidelity of probe was tested by FISH for cultured and uncultured human lymphocytes, amniocytes, chorionic villus cells, embryos, sperms, and germ cells of seminiferous tubule. Results: The human Y chromosome specific probe hybridized specifically to Y chromosome of the cells that had been tested. This probe assigned to the Yq12 region where the DYZ1 repetitive sequence is concentrated. Conclusion: We have identified a human Y chromosome specific probe that hybridized specifically to the Y chromosome by FISH for various types of uncultured as well as cultured cells. Therefore FISH technique using human Y chromosome specific probe should be useful for clinical application as a diagnostic tool for the detection of human Y chromosome.

• Journal of Microbiology and Biotechnology
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• v.27 no.10
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• pp.1778-1782
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• 2017

To identify and discriminate the bacterial species at the subspecific level, rep-PCR is a reliable genomic fingerprinting tool. Fourteen strains of bacteria were isolated from different food sources, identified as Leuconostoc citreum using 16S rRNA gene sequencing, and amplified using rep-primers (REP, ERIC, and $(GTG)_5$). Fingerprinting patterns generated bands in the range of 300-6,000 bp with REP, 150-6,000 bp with ERIC, and 200-1,700 bp with $(GTG)_5$ primers. In UPGMA dendrogram analysis, 14 strains were clustered into three clades (I, II, and III) with all the primers, thus differentiating them at the molecular level. The present study revealed the differentiation of L. citreum strains using rep-PCR.

• BMB Reports
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• v.42 no.12
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• pp.823-828
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• 2009

Techniques for analyzing protein-DNA interactions on a genome-wide scale have recently established regulatory roles for distal enhancers. However, the large sizes of higher eukaryotic genomes have made identification of these elements difficult. Information regarding sequence conservation, exon annotation and repetitive regions can be used to reduce the size of the search region. However, previously developed resources are inadequate for consolidating such information. CONVIRT is a web resource for the identification of transcription factor binding sites and also features comparative genomics. Genomic information on ortholog-independent conserved regions, exons, repeats and sequences is integrated into the virtual chromosome, and statistically over-represented single or combinations of transcription factor binding sites are sought. CONVIRT provides regulatory network analysis for several organisms with long promoter regions and permits inter-species genome alignments. CONVIRT is freely available at http://biosoft.kaist.ac.kr/convirt.

• Journal of Applied Biological Chemistry
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• v.42 no.2
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• pp.57-61
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• 1999

Seven numerically dominant 2-methyl-4-chlorophenoxyacetic acid (MCPA)-degrading bacteria were isolated from agricultural soils. The isolates utilized the herbicide MCPA as a sole carbon source, producing significant biomass in MCPA mineral medium. They exhibited diverse herbicide degradation capabilities, but most of them grew very slowly in mineral medium containing herbicide. The chromosomal DNA patterns of the isolates obtained by polymerase chain reaction amplification of repetitive extragenic palindromic sequences were distinct from each other. One isolate, SH3, which was identified as Sphingomonas species by fatty acid methyl ester analysis, was able to degrade 5 different phenoxyacetic acid herbicides within 4 days. This strain contains two plasmids, and the smaller one has a crucial role in herbicide degradation. MCPA treated into agricultural soils without indigenous MCPA-degraders persisted for a long time, but the application of the isolate SH3 resulted in rapid decline of MCPA concentration in the soil.