• Genomics & Informatics
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• v.10 no.4
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• pp.249-255
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• 2012

In this study, fosmid cloning strategies were used to assess the microbial populations in water from the International Space Station (ISS) drinking water system (henceforth referred to as Prebiocide and Tank A water samples). The goals of this study were: to compare the sensitivity of the fosmid cloning strategy with that of traditional culture-based and 16S rRNA-based approaches and to detect the widest possible spectrum of microbial populations during the water purification process. Initially, microbes could not be cultivated, and conventional PCR failed to amplify 16S rDNA fragments from these low biomass samples. Therefore, randomly primed rolling-circle amplification was used to amplify any DNA that might be present in the samples, followed by size selection by using pulsed-field gel electrophoresis. The amplified high-molecular- weight DNA from both samples was cloned into fosmid vectors. Several hundred clones were randomly selected for sequencing, followed by Blastn/Blastx searches. Sequences encoding specific genes from Burkholderia, a species abundant in the soil and groundwater, were found in both samples. Bradyrhizobium and Mesorhizobium, which belong to rhizobia, a large community of nitrogen fixers often found in association with plant roots, were present in the Prebiocide samples. Ralstonia, which is prevalent in soils with a high heavy metal content, was detected in the Tank A samples. The detection of many unidentified sequences suggests the presence of potentially novel microbial fingerprints. The bacterial diversity detected in this pilot study using a fosmid vector approach was higher than that detected by conventional 16S rRNA gene sequencing.

• Journal of Microbiology and Biotechnology
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• v.9 no.6
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• pp.744-750
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• 1999

Forty alcohol yeast strains were isolated from the main mashes (10 strains from each mash) for brewing of 4 different kinds of Korean traditional liquor (3 different types of Yakju and 1 Takju). Thirty-eight out of 40 strains were identified to be the same strain, Saccharomyces boulardii, by the Automated Bacteria, Yeast, and Fungi Identification System (Biolog Co., U.S.A.) based on the metabolic fingerprints. One strain that showed the highest ethanol production among the 38 strains in YPD medium, designated SHY 111, was selected and used for differentiating from other yeast type strains using the polymerase chain reaction (PCR). Amplified DNA, from transcribed internal spacers of SHY 111 chromosomal DNA, was found to be the same in both size and sequence as those of S. cerevisiae KCCM 11215 (formerly S. coreanus) and S. boulardii along with that of S. cerevisiae AB 972, which was used as a type strain for the yeast genome project. However, when PCR was carried out with the intron splice site primer, it resulted in the amplification of the SHY 111-specific DNA fragment which was about 200 bp in size. When PCR was carried out using the primer to test diversity of 40 isolated yeast strains, it was found that the PCR patterns were similar to each other except for the 200 bp bands derived from all the 10 strains from one Yakju, and 2 strains from another Yakju. These results suggest the strain identified as S. boulardii by the Automated Identification System to be a dominant strain for the fermentation of Korean traditional liquors.

• Journal of Plant Biotechnology
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• v.46 no.1
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• pp.1-8
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• 2019

The main goal of this study was to determine the genetic diversity among 36 grape cultivars grown in Palestine by using ISSR-polymerase chain reaction (PCR) fingerprints. Among the tested primers, 17 produced reasonable amplification products with high intensity and pattern stability. A total of 57 DNA fragments (loci) separated by electrophoresis on agarose gels were detected and they ranged in size, from 150 to 900 bp. Out of these fragments, 55 (88%) were polymorphic and 2 (3.5%) monomorphic. Our results also revealed an average of 3.1 loci per primer. A minimum of 1 and maximum of 10 DNA fragments were obtained (S-17, #820 and #841) and (S-31) primers, respectively. Therefore, the later primer (S-31) is considered to be the most powerful primer among the tested ones. The genetic distance matrix showed an average distance range of between 0.05 and 0.76. The maximum genetic distance value of 0.76 (24% similarity) was exhibited between the (Shami and Marawi.Hamadani.Adi) as well as (Bairuti and Marawi.Hamadani.Adi) genotypes. On the other hand, the lowest genetic distance of 0.05 (95% similarity) was exhibited between (Jandali.Tawel.Mofarad and Jandali. Kurawi.Mlzlz) along with (Shami.Aswad and Shami.mtartash. mlwn) genotypes. Furthermore, the UPGMA dendrogram generally clusters the grape cultivars into eight major clusters in addition to an isolated genotype. Based on these figures, the cultivars tested in this study could be characterized by large divergence at the DNA level. This is taking the assumption that our region has a very rich and varied clonal grape genetic structure.

• The Korean Journal of Mycology
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• v.24 no.2 s.77
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• pp.111-126
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• 1996

Transfer of the isolated nuclei from Lentinus edodes into protoplasts of Pleurotus florida was induced with polyethlene glycol (PEG) and $CaCl_2$. The intergeneric transfer products were classified into nuclear hybrid, heterokaryon or synkaryon, and reconstituted cell. These progenies except nuclear hybrids formed mature fruiting bodies on sawdust rice bran medium. Formation of fruit bodies was influenced by several factors such as light, temperature, nutrition and physic state of the culture media. Most of fruiting body characters were similar to those of P. florida in synkaryon and L. edodes in reconstituted cell, respectively. All these basidiocarps had clamp connections though initial heterokaryon colonies were lacking. Isozyme patterns of intergeneric progenies were quite different from those of parents. DNA polymorphisms of transfer products were also compared by random amplified polymorphic DNAs (RAPD) analysis based on polymerase chain reaction. The RAPD patterns were different from those of donor and recipient. DNA fingerprints ranged in size from 0.25 to 4.0 Kb. On the basis of RAPD, the transfer products were classified into five groups. Two synkaryon were analysed with distribution of progenies and segregation of genetic markers by random spore analyses. The genetic markers were segregated into wild type and riboflavine requiring auxotrophs.

• Microbiology and Biotechnology Letters
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• v.35 no.2
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• pp.150-157
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• 2007

The advanced bioremediation of diesel-contaminated soil through the exploration of bacterial interaction with plants was studied. A diesel-degrading rhizobacterium, Rhodococcus sp.412, and a plant species, Zea mays, having tolerant against diesel was selected. Zea mays was seeded in uncontaminated soil or diesel-contaminated soil with or without Rhodococcus sp. 412. After cultivating for 30 days, the growth of Zea mays in the contaminated soil inoculated with Rhodococcus sp. 412 was better than that in the contaminated soil without the bacterium. The residual diesel concentrations were lowered by seeding Zea mays or inoculating Rhodococctis sp. 412. These results Indicate that the simultaneous use of Zea mays and Rhodococcus sp. 412 can give beneficial effect to the remediation of oil-contaminated soil. Bacterial community was characterized using a 16S rDNA PCR and denaturing gradient gel electrophoresis (DGGE) fingerprinting method. The similarities of DGGE fingerprints were $20.8{\sim}39.9%$ between the uncontaminated soil and diesel contaminated soil. The similarities of DGGE fingerprints were $21.9%{\sim}53.6%$ between the uncontaminated soil samples, and $31.6%{\sim}50.0%$ between the diesel-contaminated soil samples. This results indicated that the structure of bacterial community was significantly influence by diesel contamination.

• Horticultural Science & Technology
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• v.32 no.6
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• pp.853-863
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• 2014

This study was carried out to construct a DNA profile database of 100 strawberry cultivars using microsatellite markers. Two hundred seventy four microsatellite primer pairs were screened with a set of 21 strawberry cultivars with different morphological traits. Twenty five primer pairs were selected because they produced reliable and reproducible fingerprints. These primer pairs were used to develop DNA profiles of 100 strawberry cultivars. Three to thirteen alleles were detected by each marker with an average of 7.50. The average polymorphism information content varied from 0.331 to 841 (average 0.706). Cluster analysis showed that the 100 cultivars were divided into 7 major groups reflecting geographic origin and pedigree information. Moreover, most of the cultivars could be discriminated by marker genotypes. These markers will be useful as a tool for the protection of plant breeders' intellectual property rights in addition to providing the means to intervene seed disputes relating to variety authentication.

• Horticultural Science & Technology
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• v.32 no.2
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• pp.210-216
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• 2014

The distribution of S-haplotypes and genetic relationships were evaluated for 47 accessions of 'Danji' radish (Raphanus sativus L. var. hortensis Baker f. gigantissimus Makino) originating from Jeju Island in South Korea. A total of 22 S-haplotype-specific SCAR markers for the S locus glycoprotein (SLG) and S receptor kinase (SRK) loci were tested, and six primer sets amplified locus-specific PCR fragments from at least one 'Danji' radish accession. S5 and S21 alleles atthe SLG locus were the most frequently distributed, and detected from 87.5% and 64.6% of the accessions, respectively. The frequency of the class-II haplotype at the SLG locus was 75%, more frequent than the class-I haplotype. The S23 allele at the SRK locus was detected from 7 accessions. Grouping of the accessions based on S-allele composition revealed three major groups, while 8 accessions showed a unique allelic composition. The genetic diversity of 47 'Danji' radishes and 1 'Gwandong' radish were also evaluated with 38 RAPD primers. A total of 312 bands were scored, and showed that 138 bands (44.2%) were monomorphic among the accessions, whereas 174 (55.8%) bands were polymorphic. Polymorphism rates ranged from 0.2 to 1.0, indicating significant variations in detecting polymorphism across RAPD primers. The genetic similarity coefficients among all pairs of the 48accessions varied from 0.62 to 0.93, and 42% of the comparisons exhibited values higher than 0.85. All the cultivars could be distinguished based on the DNA fingerprints revealed by RAPD. The comparisons between the dendrograms based on S-haplotypes and RAPDs indicate an unrelated and sporadic distribution for several accessions; however, there was a tendency for accessions with the same S-allelic composition to group into the same cluster.

• Journal of Microbiology and Biotechnology
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• v.20 no.1
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• pp.169-178
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• 2010

The microbial community structure in Thailand soils contaminated with low and high levels of arsenic was determined by denaturing gradient gel electrophoresis. Band pattern analysis indicated that the bacterial community was not significantly different in the two soils. Phylogenetic analysis obtained by excising and sequencing six bands indicated that the soils were dominated by Arthrobacter koreensis and $\beta$-Proteobacteria. Two hundred and sixty-two bacterial isolates were obtained from arsenic-contaminated soils. The majority of the As-resistant isolates were Gramnegative bacteria. MIC studies indicated that all of the tested bacteria had greater resistance to arsenate than arsenite. Some strains were capable of growing in medium containing up to 1,500 mg/l arsenite and arsenate. Correlations analysis of resistance patterns of arsenite resistance indicated that the isolated bacteria could be categorized into 13 groups, with a maximum similarity value of 100%. All strains were also evaluated for resistance to eight antibiotics. The antibiotic resistance patterns divided the strains into 100 unique groups, indicating that the strains were very diverse. Isolates from each antibiotic resistance group were characterized in more detail by using the repetitive extragenic palindromic-PCR (rep-PCR) DNA fingerprinting technique with ERIC primers. The PCR products were analyzed by agarose gel electrophoresis. The genetic relatedness of 100 bacterial fingerprints, determined by using the Pearson product-moment similarity coefficient, showed that the isolates could be divided into four clusters, with similarity values ranging from 5-99%. Although many isolates were genetically diverse, others were clonal in nature. Additionally, the arsenic-resistant isolates were examined for the presence of arsenic resistance (ars) genes by using PCR, and 30% of the isolates were found to carry an arsenate reductase encoded by the arsC gene.

• Journal of Microbiology and Biotechnology
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• v.23 no.5
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• pp.614-622
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• 2013

Daqu, a traditional fermentation starter, has been used to produce attractively flavored foods such as vinegar and Chinese liquor for thousands of years. Although Bacillus spp. are one of the dominant microorganisms in Daqu, more precise information is needed to reveal why and how Bacillus became dominant in Daqu, and next, to assess the impact of Bacillus sp. on Daqu and its derived products. We combined culture-dependent and culture-independent methods to study the ecology of Bacillus during Daqu incubation. Throughout the incubation, 67 presumptive Bacillus spp. isolates were obtained, 52 of which were confirmed by 16S rDNA sequencing. The identified organisms belonged to 8 Bacillus species: B. licheniformis, B. subtilis, B. amyloliquefaciens, B. cereus, B. circulans, B. megaterium, B. pumilus, and B. anthracis. A primer set specific for Bacillus and related genera was used in a selective PCR study, followed by a nested DGGE PCR targeting the V9 region of the 16S rDNA. Species identified from the PCR-DGGE fingerprints were related to B. licheniformis, B. subtilis, B. amyloliquefaciens, B. pumilus, B. benzoevorans, and B. foraminis. The predominant species was found to be B. licheniformis. Certain B. licheniformis strains exhibited potent antimicrobial activities. The greatest species diversity occurred at the Liangmei stage of Daqu incubation. To date, we lack sufficient knowledge of Bacillus distribution in Daqu. Elucidating the ecology of Bacillus during Daqu incubation would enable the impact of Bacillus on Daqu to be accessed, and the quality and stabilization of Daqu-derived products to be optimized.

• Journal of Microbiology and Biotechnology
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• v.18 no.12
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• pp.1966-1974
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• 2008

A typical tropical soil from the northeast of Brazil, where an important terrestrial oil field is located, was accidentally contaminated with a mixture of oil and saline production water. To study the bioremediation potential in this area, molecular methods based on PCR-DGGE were used to determine the diversity of the bacterial communities in bulk and in contaminated soils. Bacterial fingerprints revealed that the bacterial communities were affected by the presence of the mixture of oil and production water, and different profiles were observed when the contaminated soils were compared with the control. Halotolerant strains capable of degrading crude oil were also isolated from enrichment cultures obtained from the contaminated soil samples. Twenty-two strains showing these features were characterized genetically by amplified ribosomal DNA restriction analysis (ARDRA) and phenotypically by their colonial morphology and tolerance to high NaCl concentrations. Fifteen ARDRA groups were formed. Selected strains were analyzed by 16S rDNA sequencing, and Actinobacteria was identified as the main group found. Strains were also tested for their growth capability in the presence of different oil derivatives (hexane, dodecane, hexadecane, diesel, gasoline, toluene, naphthalene, o-xylene, and p-xylene) and different degradation profiles were observed. PCR products were obtained from 12 of the 15 ARDRA representatives when they were screened for the presence of the alkane hydroxylase gene (alkB). Members of the genera Rhodococcus and Gordonia were identified as predominant in the soil studied. These genera are usually implicated in oil degradation processes and, as such, the potential for bioremediation in this area can be considered as feasible.