• Journal of Life Science
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• v.22 no.2
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• pp.232-238
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• 2012

Kumjungsansung-Makgeolli$^{(R)}$ is a traditional Korean rice wine that is fermented from traditional nuruk and rice. In this study, we performed the PCR-denaturing gradient gel electrophoresis (DGGE) analysis targeting the 16S and 28S rRNA genes to characterize bacterial and fungal diversity during Makgeolli fermentation. The predominant bacteria in the PCR-DGGE profile during Makgeolli fermentation were Lactobacillus spp. (Lactobacillus curvatus, L. kisonensis, L. plantarum, L. sakei, and L. gasseri), Pediococcus spp. (P. acidilactici, P. parvulus, P. agglomerans, and P. pentosaceus), Pantoea spp. (P. agglomerans and P. ananatis), and Citrobacter freundii; these were identified on the base of analysis of 16S rRNA gene sequences. The dominant bacterium during Makgeolli fermentation was L. curvatus. The predominant fungi in PCR-DGGE profile during Makgeolli fermentation were Pichia kudriavzevii, Saccharomyces cerevisiae, Asidia idahoensis, Kluyveromyces marxianus, Saccharomycopsis fibuligera, and Torulaspora delbrueckii, and these were identified on the basis of analysis of 28S rRNA gene sequences. The dominant fungal species during Makgeolli fermentation changed from P. kudriavzevii at 0-2 days incubation to S. cerevisiae at 3-6 days incubation. This study suggests that PCR-DGGE analysis could be a suitable tool for the understanding of microbial diversity and structure during Makgeolli fermentation.

• Journal of Life Science
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• v.30 no.9
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• pp.772-782
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• 2020

Skeletal muscle differentiation or myogenesis is important to maintain muscle mass and metabolic homeostasis. Muscle-specific microRNAs (miRNAs) are known to play a critical role in skeletal myogenic differentiation. In this study, we examined the expression profiling of miRNAs during myogenic differentiation in rat L6 myoblasts using rat miRNA microarrays. We identified the upregulated expression of miR-128 as well as several well-known myogenic miRNAs, including miR-1, miR-133b, and miR-206. We additionally confirmed the increased expression of miR-128 observed on microarray through quantitative real-time PCR (qRT-PCR), which showed similarly upregulated expression of both primary miR-128 and mature miR-128, consistent with the microarray findings. Furthermore, transfection of miR-128 into rat L6 myoblasts induced gene expression of myogenic markers such as muscle creatine kinase (MCK), myogenin, and myosin heavy chain (MHC). Protein expression of MHC was increased as well. Inhibition of miR-128 by inhibitory peptide nucleic acids (PNAs) blocked the expression of those myogenic markers. In addition, the transfection of miR-128 into rat L6 myoblasts enhanced the phosphorylation of Erk and Akt proteins stimulated by insulin, while simultaneously reversing the inhibited phosphorylation of Erk and Akt due to insulin resistance. These findings suggest that miR-128 may play important roles in myogenic differentiation and insulin signaling.

• The Journal of the Korean Society for Microbiology
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• v.35 no.4
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• pp.283-288
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• 2000

The PCR primer set JW21-JW22 of Weiss et al. (19), which was reported to amplify a 139-bp fragment of the l6S rRNA gene of Helicobacter pylori, has been recently used for the detection of H. pylori in clinical specimens. However, when we applied JW21-JW22 PCR to other members of the genus Helicobacter and unrelated microorganisms, all of these bacteria produced a 139-bp PCR product. Therefore, we designed a novel primer set, HPU185-HPL826, which produced a 642-bp amplicon of the l6S rRNA gene of H. pylori. Then we further examined the specificity of the novel PCR assay using Southern blot hybridization with an internal probe, HPP225. The PCR assay described in this study was shown to be highly sensitive and specific only to the H. pylori 16S rRNA gene sequences.

• Journal of the Korean Chemical Society
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• v.42 no.2
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• pp.209-213
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• 1998

The higher order structure of Pseudomonas alcaligenes 5S rRNA has been investigated by using ($\eta^{6}$-mesitylene) manganese (Ⅰ) tricarbonyl hexafluorophosphate[MTH-Mn (Ⅰ)], dimethylpyrocarbonate, potassium permanganate as chemical probes. The sequences cleaved strongly by MTH-Mn (Ⅰ) on the tertiary structure of the 5S rRNA are $G_{12}AUGG_{16}$ of loop a, $G_{51}AAGUGAAGC_{60}$ of the region b-C, $U_{65}-AGCG_{69}$. of the region B-a, and $G_{72}AUGG_{76}$ of loop d. Based on such cleavage patterns of 5S rRNA by MTH-Mn(Ⅰ) and other chemical probes, we presume that the sequences strongly cleaved form pocket-like structure as in the the corner of L structure of $tRNA^{Phe}$. We also presume that the region b-C and loop d together play a role of hinge in forming the pocket-like structure in the 5S rRNA.

• Korean Journal of Environmental Biology
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• v.21 no.1
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• pp.77-85
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• 2003

We have examined the 16S-23S rRNA intergenic spacer regions (ISR) of Vibrio fluvialis. ISRs were PCR amplified, cloned into a plasmid vector and then sequenced. As results of ISR nucleotide sequence analysis, total of 6 clones were isolated depending on the size. The clones were different in both the number and the composition of the tRNA genes, and were designated ISR-A, ISR-E, ISR-El, ISR-lA, ISR-EKV, ISR-EKAV. ISR-A contains $tRNA^{Ala}$; ISR-lA, $tRNA^{Ile}$-$tRNA^{Ala}$; ISR-EKV, $tRNA^{GIu}$-$tRNA^{Lys}$-$tRNA^{Val}$;ISE-EKAV, $tRNA^{GIu}$-$tRNA^{Lys}$-$tRNA^{Ala}$-$tRNA^{Val}$; ISR -E and E1, $tRNA^{GIu}$ clusters. ISR-EKV was shown to be a minor type out of the six ISR types and showed a very limited homology between ISR-EKV from V, fluvialis and ISRa from other Vibrio species. Therefore ISR-EKV sequence was used to design species-specific primers to detect V, fiuvialis from other Vibrio species by PCR reaction. The specificity of the primers was examined using genomic DNA of other Vibrios as templates for PCR reaction. The result showed that PCR can be a useful method to detect V. fluvialis among Vibrio species in a single PCR reaction.

• Korean Journal of Microbiology
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• v.44 no.3
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• pp.251-257
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• 2008

Four potential microcystin-producing cyanobacteria were isolated from large reservoirs which act as sources of drinking water supply in Korea. Strain DC-2, YD-l, and YD-6 were closely related to Microcystis aeruginosa based on the analysis of l6S rRNA gene and mcyA gene sequences. mcyA gene sequence of YDS2-3, isolated from Yongdam Reservoir, was closed to that of M. aeruginosa, whereas l6S rRNA gene sequence was not related to the known sequences of microcystin-producing cyanobacteria indicating this strain can be a novel cyanobacterium belonging to the genus Microcystis. When mcyA gene sequences of isolated cyanobacteria were compared with the mcyA gene sequence library of two reservoirs, the sequence of DC-2 matched with the dominant ones.

• Journal of Microbiology and Biotechnology
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• v.17 no.11
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• pp.1743-1750
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• 2007

Two Gram-negative, nonmotile, coccobacilli, SW-$3^T$ and SW-$100^T$, were isolated from sea water of the Yellow Sea in Korea. Strains SW-$3^T$ and SW-$100^T$ contained ubiquinone-9 (Q-9) as the predominant respiratory lipoquinone and $C_{18:1}\;{\omega}9c$ and $C_{16:0}$ as the major fatty acids. The DNA G+C contents of strains SW-$3^T$ and SW- $100^T$ were 44.1 mol% and 41.9 mol%, respectively. A neighbor-joining tree based on l6S rRNA gene sequences showed that the two isolates fell within the evolutionary radiation enclosed by the genus Acinetobacter. Strains SW-$3^T$ and SW-$100^T$ exhibited a l6S rRNA gene similarity value of 95.7% and a mean DNA-DNA relatedness level of 9.2%. Strain SW-$3^T$ exhibited l6S rRNA gene sequence similarity levels of 93.5-96.9% to the validly described Acinetobacter species and fifteen Acinetobacter genomic species. Strain SW-$100^T$ exhibited l6S rRNA gene sequence similarity levels of less than 97.0% to the other Acinetobacter species except Acinetobacter towneri DSM $14962^T$ (98.0% similarity). Strains SW-$3^T$ and SW-$100^T$ exhibited mean levels of DNA-DNA relatedness of 7.3-l6.7% to the type strains of some phylogenetically related Acinetobacter species. On the basis of phenotypic, phylogenetic, and genetic data, strains SW-$3^T$ and SW-$100^T$ were classified in the genus Acinetobacter as two distinct novel species, for which the names Acinetobacter marinus sp. novo (type strain SW-$3^T$=KCTC $12259^T$=DSM $16312^T$) and Acinetobacter seohaensis sp. novo (type strain SW-$100^T$=KCTC $12260^T$=DSM $16313^T$) are proposed, respectively.

• Korean Journal of Microbiology
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• v.47 no.4
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• pp.289-294
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• 2011

We investigated the cold shock sensitivity of DEAD-box RNA helicase gene deleted strains of in Bacillus subtilis CU1065. To understand cold shock effects, cells were cultivated at $37^{\circ}C$ to log phase ($O.D_{600}$=0.5-0.6) and then temperature was shifted to $15^{\circ}C$. Cold shock slow down the growth rate of wild type and deleted strains of DEAD-box RNA helicase gene (ydbR, yfmL, yqfR, deaD). The growth rate of ydbR deleted strain is 5 times severely reduced compared to that of wild type strain (CU1065). But the growth rate of other three (yfmL, yqfR, deaD) deleted strains is nearly equal to the growth rate of wild type. Compared to $37^{\circ}C$, the amount of ydbR and yqfR mRNA transcripts are increased at the growth temperature of $15^{\circ}C$. On the other hands the mRNA transcripts of yfmL and deaD are not changed at both conditions of $37^{\circ}C$ and $15^{\circ}C$. Upon cold shock treatment ydbR mRNA transcript is clearly increased. After treatment of rifampicin (bacteria transcription inhibitor) the amount of ydbR mRNA was measured. Temperature shift from $37^{\circ}C$ to $15^{\circ}C$ and rifampicin treatment showed slowly decay of ydbR mRNA. But at $37^{\circ}C$ and rifampicin treatment ydbR mRNA is rapidly reduced. These results showed that cold shock induction of ydbR mRNA resulted from the stability of ydbR mRNA and not from the transcription induction of ydbR. In relation to these results, we found the cold box element of csp (cold shock protein gene) in 5' untranslated region of ydbR gene. Cold shock induction of ydbR is caused by the stability of ydbR mRNA like the stability of csp mRNA.

• Clinical and Experimental Pediatrics
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• v.53 no.2
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• pp.178-183
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• 2010

Purpose : The aim of this study was to identify mutations associated with macrolide resistance in Mycoplasma pneumoniae (MP) and to establish a cultural method to determine antimicrobial susceptibility. Methods : Nasopharyngeal aspirates (NPAs) were collected from 62 children diagnosed with MP pneumonia by a serologic method or polymerase chain reaction. The 23S rRNA and L4 ribosomal protein genes of MP were amplified and sequenced. To identify mutations in these 2 genes, their nucleotide sequences were compared to those of the reference strain M129. MP cultivation was carried out for 32 (28 frozen and 5 refrigerated) NPAs and M129 strain using Chanock's glucose broth and agar plate in a 5% $CO_2$ incubator at $37^{\circ}C$ and examined at 2-3 day intervals for 6 weeks. Results : Among the 62 specimens, 17 had M144V mutations in ribosomal protein L4. The A2064G mutation was observed in 1 specimen; its 23S rRNA gene was successfully sequenced. Culture for MP was successful from the M129 strain and 2 of the 5 NPAs that were refrigerated for no longer than 3 days. However, MP did not grow from the 28 NPAs that were kept frozen at $-80^{\circ}C$ since 2003. Conclusion : We found the M144V mutation of L4 protein to be common and that of domain V of 23S rRNA gene was relatively rare among MP. Studies on the prevalence of macrolide-resistant MP and the relationship between the mutations of 23S rRNA gene and ribosomal protein L4 will aid in understanding the mechanism of macrolide resistance in MP.

• Fisheries and Aquatic Sciences
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• v.3 no.1
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• pp.64-70
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• 2000

DNA probes for the l6S rRNA have been designed for the detection of Edwardsiella tarda. In order to accomplish this purpose, the l6S rRNA gene from E. tarda has been cloned and sequenced. Two highly feasible oligonucleotide probe sites have been determined by the database analysis programs presented by PCGENE and BLAST. These two probes have been evaluated by slot blot hybridization analysis. Hetero- and homo-trimeric templates have been synthesized using these two probe sites. The templates have been further multimerized by PCR to generate between 150 and 300 bp long DIG-11-dUTP labeled probes. Unlike 3' end labeled oligonucleotide probes or templates, multimerized probes showed no cross­hybridization in the given experimental condition. Furthermore, a significant increase in sensitivity has been observed with these probes. This method, we presented here, may be useful for the designing of probes for the detection of other fish pathogenic microorganisms also.