• Animal Bioscience
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• v.36 no.4
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• pp.654-670
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• 2023

Objective: This study aimed to develop loop-mediated isothermal amplification (LAMP) combined with lateral flow dipstick (LFD) and compare it with LAMP-AGE, polymerase chain reaction (PCR), and standard Salmonella culture as reference methods for detecting Salmonella contamination in animal products and animal production environmental samples. Methods: The SalInvA01 primer, derived from the InvA gene and designed as a new probe for LFD detection, was used in developing this study. Adjusting for optimal conditions by temperature, time, and reagent concentration includes evaluating the specificity and limit of detection. The sampling of 120 animal product samples and 350 animal production environmental samples was determined by LAMP-LFD, comparing LAMP-AGE, PCR, and the culture method. Results: Salmonella was amplified using optimal conditions for the LAMP reaction and a DNA probe for LFD at 63℃ for 60 minutes. The specificity test revealed no cross-reactivity with other microorganisms. The limit of detection of LAMP-LFD in pure culture was 3×10² CFU/mL (6 CFU/reaction) and 9.01 pg/μL in genomic DNA. The limit of detection of the LAMP-LFD using artificially inoculated in minced chicken samples with 5 hours of pre-enrichment was 3.4×10⁴ CFU/mL (680 CFU/reaction). For 120 animal product samples, Salmonella was detected by the culture method, LAMP-LFD, LAMP-AGE, and PCR in 10/120 (8.3%). In three hundred fifty animal production environmental samples, Salmonella was detected in 91/350 (26%) by the culture method, equivalent to the detection rates of LAMP-LFD and LAMP-AGE, while PCR achieved 86/350 (24.6%). When comparing sensitivity, specificity, positive predictive value, and accuracy, LAMP-LFD showed the best results at 100%, 95.7%, 86.3%, and 96.6%, respectively. For Kappa index of LAMP-LFD, indicated nearly perfect agreement with culture method. Conclusion: The LAMP-LFD Salmonella detection, which used InvA gene, was highly specific, sensitive, and convenient for identifying Salmonella. Furthermore, this method could be used for Salmonella monitoring and primary screening in animal products and animal production environmental samples.

• 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.

• Animal Bioscience
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• v.36 no.7
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• pp.991-1002
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• 2023

Objective: This study aimed to elucidate the underlying gene regions responsible for productive, phenotypic or adaptive traits in different ecological types of Tibetan sheep and the discovery of important genes encoding valuable traits. Methods: We used whole-genome resequencing to explore the genetic relationships, phylogenetic tree, and population genetic structure analysis. In addition, we identified 28 representative Tibetan sheep single-nucleotide polymorphisms (SNPs) and genomic selective sweep regions with different traits in Tibetan sheep by fixation index (Fst) and the nucleotide diversity (θπ) ratio. Results: The genetic relationships analysis showed that each breed partitioned into its own clades and had close genetic relationships. We also identified many potential breed-specific selective sweep regions, including genes associated with hypoxic adaptability (MTOR, TRHDE, PDK1, PTPN9, TMTC2, SOX9, EPAS1, PDGFD, SOCS3, TGFBR3), coat color (MITF, MC1R, ERCC2, TCF25, ITCH, TYR, RALY, KIT), wool traits (COL4A2, ERC2, NOTCH2, ROCK1, FGF5, SOX9), and horn phenotypes (RXFP2). In particular, a horn-related gene, RXFP2, showed the four most significantly associated SNP loci (g. 29481646 A>G, g. 29469024 T>C, g. 29462010 C>T, g. 29461968 C>T) and haplotypes. Conclusion: This finding demonstrates the potential for genetic markers in future molecular breeding programs to improve selection for horn phenotypes. The results will facilitate the understanding of the genetic basis of production and adaptive unique traits in Chinese indigenous Tibetan sheep taxa and offer a reference for the molecular breeding of Tibetan sheep.

• Korean Journal of Environmental Biology
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• v.41 no.1
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• pp.1-13
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• 2023

Broad bean wilt virus 2 (BBWV2) is a species in the genus Fabavirus and family Secoviridae, which is transmitted by aphids and has a wide host range. The BBWV2 genome is composed of two single-stranded, positive-sense RNAs, RNA-1 and RNA-2. The representative symptoms of BBWV2 are mosaic, mottle, vein clearing, wilt, and stunting on leaves, and these symptoms cause economic damage to various crops. In 2019, Perilla fructescens leaves with mosaic and yellowing symptoms were found in Geumsan, South Korea. Reverse-transcription polymerase chain reaction (RT-PCR) was performed with specific primers for 10 reported viruses, including BBWV2, to identify the causal virus, and the results were positive for BBWV2. To characterize a BBWV2 isolate (BBWV2-GS-PF) from symptomatic P. fructescens, genetic analysis and pathogenicity tests were performed. The complete genomic sequences of RNA-1 and RNA-2 of BBWV2-GS-PF were phylogenetically distant to the previously reported BBWV2 isolates, with relatively low nucleotide sequence similarities of 76-80%. In the pathogenicity test, unlike most BBWV2 isolates with mild mosaic or mosaic symptoms in peppers, the BBWV2-GS-PF isolate showed typical ring spot symptoms. Considering these results, the BBWV2-GS-PF isolate from P. fructescens could be classified as a new strain of BBWV2.

• Korean Journal of Animal Reproduction
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• v.25 no.3
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• pp.237-242
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• 2001

We recently demonstrated that both linear- and supercoil-form B1/B2 SINE (short interspersed elements) sequences could increase an integration frequency of a reporter gene in preimplatation mouse embryos. In those reports, when either a control or SINE-flanked DNA was separately applied to microinjection, the proportions of $\beta$-gal positives were 16% and 63%, respectively, in linear-form DNA, and 6% and 25%, respectively, in circular-form DNA. Here, we examined the contribution of a circular-form DNA moiety to integration frequency by using a mixed-farm (linear and circular-form) DNA in microinjection. When examined in the blastocyst stage, the proportion of $\beta$-gal-positive embryos was 17.3% and 46.6% in control and SINE-flanked DNA, respectively. These results suggest that there is little contribution of circular-form DNA moiety to the resultant integration frequency, and that the majority of the integration events are mediated through a linear conformation of vector DNA. In addition, some clues on integration process could be obtained from the analysis of microinjection results.

• Korean Journal of Audiology
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• v.23 no.1
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• pp.20-26
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• 2019

Background and Objectives: Autosomal recessive non-syndromic hearing loss (ARNSHL) with genetic origin is common (1/2000 births). ARNSHL can be associated with mutations in gap junction protein beta 2 (GJB2). To this end, this cohort investigation aimed to find the contribution of GJB2 gene mutations with the genotype-phenotype correlations in 45 ARNSHL cases in the Kurdish population. Subjects and Methods: Genomic DNA was extracted from a total of 45 ARNSHL families. The linkage analysis with 3 short tandem repeat markers linked to GJB2 was performed on 45 ARNSHL families. Only 9 of these families were linked to the DFNB1 locus. All the 45 families who took part were sequenced for confirmation linkage analysis (to perform a large project). Results: A total of three different mutations were determined. Two of which [c.35delG and c.-23+1G>A (IVS1+1G>A)] were previously reported but (c.299-300delAT) mutation was novel in the Kurdish population. The homozygous pathogenic mutations of GJB2 gene was observed in nine out of the 45 families (20%), also heterozygous genotype (c.35delG/N)+(c.-23+1G>A/c.-23+1G>A) were observed in 4/45 families (8.8%). The degree of hearing loss (HL) in patients with other mutations was less severe than patients with c.35delG homozygous mutation (p<0.001). Conclusions: Our data suggest that GJB2 mutations constitute 20% of the etiology of ARNSHL in Iran; moreover, the c.35delG mutation is the most common HL cause in the Kurdish population. Therefore, these mutations should be included in the molecular testing of HL in this population.

• Journal of agriculture & life science
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• v.50 no.3
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• pp.99-117
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• 2016

Data on primal cuts were collected from 1,829 steers of Hanwoo progeny testing programs, between 2010 and 2015 for the ssGWAS. SNP data were analyzed by using Illumina Bovine 50K Beadchip. The SNP data that matches with phenotype data was 674 animals. As a first step, the genomic estimated breeding value(GEBV) of the loin and rib cuts were estimated, which was used in the estimation of SNP marker effects and their variances related to the traits. Then, the estimated variance explained by each marker was expressed as a proportion to the total genetic variance. Finally, the SNP loci and their significance to any possible QTL were examined. Among the 20 best SNP loci explaining a larger proportion of SNP variance to the total genetic variance for tender loin yield, the region between 12,812,193 ~ 12,922,313bp on BTA 10 harbored a cluster of SNPs that explained about 7.32 to 7.34% of the total genetic variance. For strip loin yield, a peak for higher effects for multiple SNPs was found in BTA24, between 38,158,543 and 38,347,278bp distances, which explained about 8.36 to 8.56% of the observed variance for this trait. For loin yield had relatively smaller effects in terms of the total genetic variance. Therefore, loin yield might be affected by a few loci with moderate effects and many other loci with smaller effects across the genome.

• Journal of Periodontal and Implant Science
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• v.53 no.3
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• pp.233-244
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• 2023

Purpose: An implant-supported prosthesis consists of an implant fixture, an abutment, an internal screw that connects the abutment to the implant fixture, and the upper prosthesis. Numerous studies have investigated the microorganisms present on the implant surface, surrounding tissues, and the subgingival microflora associated with peri-implantitis. However, there is limited information regarding the microbiome within the internal screw space. In this study, microbial samples were collected from the supragingival surfaces of natural teeth, the peri-implant sulcus, and the implant-abutment screw hole, in order to characterize the microbiome of the internal screw space in healthy subjects. Methods: Samples were obtained from the supragingival region of natural teeth, the peri-implant sulcus, and the implant screw hole in 20 healthy subjects. DNA was extracted, and the V3-V4 region of the 16S ribosomal RNA was sequenced for microbiome analysis. Alpha diversity, beta diversity, linear discriminant analysis effect size (LEfSe), and network analysis were employed to compare the characteristics of the microbiomes. Results: We observed significant differences in beta diversity among the samples. Upon analyzing the significant taxa using LEfSe, the microbial composition of the implant-abutment screw hole's microbiome was found to be similar to that of the other sampling sites' microbiomes. Moreover, the microbiome network analysis revealed a unique network complexity in samples obtained from the implant screw hole compared to those from the other sampling sites. Conclusions: The bacterial composition of the biofilm collected from the implant-abutment screw hole exhibited significant differences compared to the supra-structure of the implant. Therefore, long-term monitoring and management of not only the peri-implant tissue but also the implant screw are necessary.

• Analytical Science and Technology
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• v.21 no.4
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• pp.338-343
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• 2008

Extraction of DNA from skeletal material is of great importance in the identification of human remains, but is particularly difficult because the high amount of microbial DNA was often co-extracted with human bone DNA. We found that a phenol/chloroform extraction, followed by ultrafiltration, and cleanup by via the $QIAquick^{(R)}$ PCR purification kit yields higher amounts of human genomic DNA compared with extraction by the column affinity $method^{(R)}$ alone. Ultrafiltration extraction of human DNA from ten exhumed bone samples yielded $0.041-1.120ng/{\mu}L$ DNA (mean = $0.498ng/{\mu}L$ DNA), and purification using the column affinity resulted in $0.016-0.064ng/{\mu}L$ DNA (mean = $0.034ng/{\mu}L$ DNA). Although the STR genotyping by the column affinity method was partially successful, all DNA samples by the ultrafiltration method produced full profiles from the multiplex PCR. The efficiency of STR genotyping was in accordance with the amounts of the human DNA extracted.

• Korean Journal of Poultry Science
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• v.20 no.4
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• pp.177-188
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• 1993

This study was performed to find out the reasonable sexing methods In the chicken, obtain the basic information for the mechanisms related to chicken sexual differentiation and identify the genes which known to involved in chicken sex differentiation. The chromosome analysis of chicken embryonic fibroblast was a simple method to determine sex of chicken by means of Z and W chromosome identification. The bands of female chicken genomic DNA digested with Xho Ⅰ and Eco RI restriction endonuclease showed to be useful in direct sex determination and these repetitive sequences of Xho Ⅰ and Eco RI families were proposed to be very homologous in their sequences by colony hybridization analysis. Seven of 150 random primers were selected to amplify the W chromosome-specific band by using arbitrary primed PCR and three of them were useful to identify the sex of chicken. To identify the sex differentiation genes in the chicken, PCR for the amplification of ZFY and SRY sequences was performed. ZFY and SRY sequences were amplified successfully in the chicken genome, implying that chicken genome might have the sex-related conserved sequences similar to mammalian ones. The PCR products of ZFY amplification were the same in both sexes, suggesting that these sequences may be located on autosome or Z chromosome. The profile of PCR amplification for SRY sequences showed variation between sexes, but this result was not enough to specify whether the SRY gene in chicken is on the autosome or sex chromosome.