• Title/Summary/Keyword: DNA-based testing

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Genotoxicity Assessment of Erythritol by Using Short-term Assay

  • Chung, Young-Shin;Lee, Michael
    • Toxicological Research
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    • v.29 no.4
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    • pp.249-255
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    • 2013
  • Erythritol is a sugar alcohol that is widely used as a natural sugar substitute. Thus, the safety of its usage is very important. In the present study, short-term genotoxicity assays were conducted to evaluate the potential genotoxic effects of erythritol. According to the OECD test guidelines, the maximum test dose was 5,000 ${\mu}g$/plate in bacterial reverse mutation tests, 5,000 ${\mu}g/ml$ in cell-based assays, and 5,000 mg/kg for in vivo testing. An Ames test did not reveal any positive results. No clastogenicity was observed in a chromosomal aberration test with CHL cells or an in vitro micronucleus test with L5178Y $tk^{+/-}$ cells. Erythritol induced a marginal increase of DNA damage at two high doses by 24 hr of exposure in a comet assay using L5178Y $tk^{+/-}$ cells. Additionally, in vivo micronucleus tests clearly demonstrated that oral administration of erythritol did not induce micronuclei formation of the bone marrow cells of male ICR mice. Taken together, our results indicate that erythritol is not mutagenic to bacterial cells and does not cause chromosomal damage in mammalian cells either in vitro or in vivo.

The Usefulness of Cervicovaginal Cytology as a Primary Screening Test (일차 선별검사로서 자궁경부 세포진 검사의 유용성)

  • Park, Jae-Hong;Ha, Seung-Yeon;Cho, Hyun-Yee;Chung, Dong-Hae;Kim, Na-Rae;Park, Sang-Hui
    • The Korean Journal of Cytopathology
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    • v.19 no.2
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    • pp.107-110
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    • 2008
  • We evaluated the usefulness of cervicovaginal cytology as a primary screening test by analyzing the cytologic and histological diagnoses of 2,254 women. Cervicovaginal cytology had 93.0% sensitivity, 86.1% specificity, 88.2% positive predictive value, and 91.7% of negative predictive value. Cervicovaginal cytology as a primary screening test showed much higher specificity but slightly lower sensitivity than HPV DNA testing. However, the sensitivity of cervicovaginal cytology will be improved continuously due to the development of liquid-based cytology. We regard cervicovaginal cytology as a good primary screening test for cervical intraepithelial neoplasia or carcinoma.

Development of real-time PCR for rapid detection of Mycobacterium bovis DNA in cattle lymph nodes and differentiation of M. bovis and M. tuberculosis (소 림프절에서 Mycobacterium bovis DNA의 신속 검출과 M. bovis와 M. tuberculosis 감별을 위한 real-time PCR 개발)

  • Koh, Ba-Ra-Da;Jang, Young-Boo;Ku, Bok-Kyung;Cho, Ho-Seong;Bae, Seong-Yeol;Na, Ho-Myung;Park, Seong-Do;Kim, Yong-Hwan;Mun, Yong-Un
    • Korean Journal of Veterinary Service
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    • v.34 no.4
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    • pp.321-331
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    • 2011
  • Mycobacterium bovis, a member of the M. tuberculosis complex (MTC), is the causative agent of bovine tuberculosis. Detection of M. bovis and M. tuberculosis using conventional culture- and biochemical-based assays is time-consuming. Therefore, a simple and sensitive molecular assay for rapid detection would be of great help in specific situations such as faster diagnosis of bovine tuberculosis (bTB) infection in the abattoirs. We developed a novel multiplex real-time PCR assay which was applied directly to biological samples with evidence of bTB and it was allowed to differentiate between M. bovis and M. tuberculosis. The primers and TaqMan probes were designed to target the IS1081 gene, the multi-copy insertion element in the MTC and the 12.7-kb fragment which presents in M. tuberculosis, not in the M. bovis genome. The assay was optimized and validated by testing 10 species of mycobacteria including M. bovis and M. tuberculosis, and 10 other bacterial species such as Escherichia coli, and cattle lymph nodes (n=113). The tests identified 96.4% (27/28) as M. bovis from the MTC-positive bTB samples using conventional PCR for specific insertion elements IS1081. And MTC-negative bTB samples (n=85) were tested using conventional PCR and the real-time PCR. When comparative analyses were conducted on all bovine samples, using conventional PCR as the gold standard, the relative accuracy of real-time PCR was 99.1%, the relative specificity was 100%, and the agreement quotient (kappa) was 0.976. The detection limits of the real-time PCR assays for M. bovis and M. tuberculosis genomic DNA were 10 fg and 0.1 pg per PCR reaction, respectively. Consequently, this multiplex real-time PCR assay is a useful diagnotic tool for the identification of MTC and differentiation of M. bovis and M. tuberculosis, as well as the epidemiologic surveillance of animals slaughtered in abattoir.

Effects of SNP Markers of the Apolipoprotein E (APOE) Gene on Meat Quantity and Quality Traits in Korean Cattle (한우 아포지단백질 E (APOE) 유전자의 SNP Marker가 육량 및 육질형질에 미치는 영향)

  • Shin, Ki-Hyun;Shin, Sung-Chul;Chung, Ku-Young;Chung, Eui-Ryong
    • Food Science of Animal Resources
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    • v.29 no.1
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    • pp.108-113
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    • 2009
  • Apolipoprotein E (APOE) is a plasma lipoprotein in mammals and plays an important role in the transport and metabolism of lipids such as phospholipids and triglycerides. Therefore, the APOE gene could be a candidate gene controlling lipid metabolism in beef cattle. This study was performed to identify single nucleotide polymorphisms (SNP) in the APOE gene and to investigate the effects of SNP genotype on the carcass traits such as meat quantity and quality in Korean cattle. For PCR amplification, pooled DNA made from unrelated 60 individuals was prepared and primer pairs were designed based on the cDNA sequence of exon 4 region of the bovine APOE gene. A SNP was identified at position 2034 (T/C substitution) of the exon 4 region in the APOE gene. PCR-RFLP procedure with restriction enzyme ACC I was developed for determining the SNP genotype for each of a total of 309 animals with pedigree information and performance records through the national progeny testing program. The frequencies of the genotypes TT, TC and CC were 10.9, 46.9 and 42.2%. Gene frequencies were 0.344 for T allele and 0.656 for C allele. The g.2034T>C SNP genotype showed a significant effect (p<0.05) on dressing percentage and meat color, respectively. Animals with the TT genotype showed higher dressing percentage than those with the CC genotype, and TT genotype had desirable meat color compared with CC genotype. These results suggest that the g.2034T>C SNP genotype of the APOE gene may be useful as a DNA marker for meat quantity index and dressing percentage in Korean cattle.

Plant Growth-Promoting Potential of Endophytic Bacteria Isolated from Roots of Coastal Sand Dune Plants

  • Shin, Dong-Sung;Park, Myung-Soo;Jung, Se-Ra;Lee, Myoung-Sook;Lee, Kang-Hyun;Bae, Kyung-Sook;Kim, Seung-Bum
    • Journal of Microbiology and Biotechnology
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    • v.17 no.8
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    • pp.1361-1368
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    • 2007
  • Endophytic bacteria associated with the roots of coastal sand dune plants were isolated, taxonomically characterized, and tested for their plant growth-promoting activities. Ninety-one endophytic bacterial isolates were collected and assigned to 17 different genera of 6 major bacterial phyla based on partial 16S rDNA sequence analyses. Gammaproteobacteria represented the majority of the isolates (65.9%), and members of Pseudomonas constituted 49.5% of the total isolates. When testing for antagonism towards plant pathogenic fungi, 25 strains were antagonistic towards Rhizoctonia solani, 57 strains were antagonistic towards Pythium ultimum, 53 strains were antagonistic towards Fusarium oxysporum, and 41 strains were antagonistic towards Botrytis cinerea. Seven strains were shown to produce indole acetic acid (IAA), 33 to produce siderophores, 23 to produce protease, 37 to produce pectinase, and 38 to produce chitinase. The broadest spectra of activities were observed among the Pseudomonas strains, indicating outstanding plant growth-promoting potential. The isolates from C. kobomugi and M. sibirica also exhibited good plant growth-promoting potential. The correlations among individual plant growth-promoting activities were examined using phi coefficients, and the resulting data indicated that the production of protease, pectinase, chitinase, and siderophores was highly related.

Genotype Distribution of Human Papillomavirus in Women with Abnormal Cervical Cytology in an Esophageal Carcinoma High Incidence Area of China

  • Mai, Rui-Qin;Huang, Bo;Shen, Ling;Zhang, Guo-Hong;Hong, Liang-Li;Cai, Ying-Mu
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.12
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    • pp.4945-4950
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    • 2014
  • Infection with human papillomavirus (HPV) could affect genesis of both cervical and esophageal cancers. The type-specific distribution of HPV in cervical cytology abnormalities of women has remained unclear in Shantou, an esophageal cancer high-incidence area of China. Data from 22,617 women who were subjected to cervical HPV DNA testing with simultaneous cervical cytological examination during 2009-2013 were therefore here retrospectively evaluated in a hospital-based study. Overall, 16.2% (3,584/22,114)of women with normal cytology were HR-HPV positive, with HPV-52 (4.07%) as the most common type followed by -16 (3.63%), and -58 (2.46%). Prevalence of HR-HPV was 50.3% (253/503) in women with cervical cytological abnormalities, of which in ASC-H 71.4%, ASC-US 39.1%, HSIL 80.3% and LSIL 73.7%. HPV-58 (14.12%) was the most common type for all cervical cytological abnormalities, followed by HPV-16 (13.72%), and -52 (12.72%), while the more common HPV-16 type in ASC-H (42.9%) and HSIL (36.1%), HPV-52 and -58 were the most common types for ASC-US (10.3%) and LSIL (25%), respectively. Multiple HPV co-infections were identified in 33.2% (84/253) cytology abnormalities with positive HR-HPV, and the highest prevalence of HPV-58/16 combination in HSIL (28.6%, 6/21) was observed. Our data indicated a relative high prevalence of HPV-58 and -52 in women with cervical cytological abnormalities, which should be considered in the development of next-generation vaccines for Shantou.

Noninvasive prenatal test for the pregnancy with Turner syndrome mosaicism 45, X/47, XXX: A case report

  • Kim, Ji Hye;Lee, Gun Ho;Cha, Dong Hyun;Cho, Eun-Hae;Jung, Yong Wook
    • Journal of Genetic Medicine
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    • v.12 no.2
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    • pp.118-122
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    • 2015
  • Noninvasive prenatal test (NIPT) is a novel screening method for the diagnosis of fetal chromosomal aneuploidies. NIPT is based on technology that detects cell-free fetal DNA in maternal plasma and analyzes it with massively parallel sequencing technology to determine whether the fetus is at risk of trisomy 21, trisomy 18, trisomy 13 or sex chromosome abnormalities (SCAs). NIPT has been reported to have sensitivity of 99% and a false positive rate of less than 1% for detecting trisomy 21 and trisomy 18. Although extension of the application of NIPT to other SCAs has been attempted, there are concerns in extending NIPT to SCAs because of maternal or fetal mosaicism, undetected maternal SCAs, and multiple pregnancies. Recently, we assessed a pregnancy with the rare Turner syndrome mosaicism 45, X/47, XXX, which was reported as 45, X with NIPT. We present the case here and briefly review the current literatures on NIPT in testing for fetal monosomy X. To the best of our knowledge, this is the first report of the 45, X/47, XXX mosaicism in Korea to be reported as 45, X by NIPT with whole genome sequencing. This case report will provide valuable information for counseling women who want to undergo NIPT.

Screening and Characterization of Lactic Acid Bacteria Strains with Anti-inflammatory Activities through in vitro and Caenorhabditis elegans Model Testing

  • Lee, Hye Kyoung;Choi, Sun-Hae;Lee, Cho Rong;Lee, Sun Hee;Park, Mi Ri;Kim, Younghoon;Lee, Myung-Ki;Kim, Geun-Bae
    • Food Science of Animal Resources
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    • v.35 no.1
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    • pp.91-100
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    • 2015
  • The present study was conducted to screen candidate probiotic strains for anti-inflammatory activity. Initially, a nitric oxide (NO) assay was used to test selected candidate probiotic strains for anti-inflammatory activity in cultures of the murine macrophage cell line, RAW 264.7. Then, the in vitro probiotic properties of the strains, including bile tolerance, acid resistance, and growth in skim milk media, were investigated. We also performed an in vitro hydrophobicity test and an intestinal adhesion assay using Caenorhabditis elegans as a surrogate in vivo model. From our screening, we obtained 4 probiotic candidate lactic acid bacteria (LAB) strains based on their anti-inflammatory activity in lipopolysaccharide (LPS)-stimulated RAW 264.7 cell cultures and the results of the in vitro and in vivo probiotic property assessments. Molecular characterization using 16S rDNA sequencing analysis identified the 4 LAB strains as Lactobacillus plantarum. The selected L. plantarum strains (CAU1054, CAU1055, CAU1064, and CAU1106) were found to possess desirable in vitro and in vivo probiotic properties, and these strains are good candidates for further investigations in animal models and human clinical studies to elucidate the mechanisms underlying their anti-inflammatory activities.

Identification of Pork Contamination in Meatballs of Indonesia Local Market Using Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) Analysis

  • Erwanto, Yuny;Abidin, Mohammad Zainal;Muslim, Eko Yasin Prasetyo;Sugiyono, Sugiyono;Rohman, Abdul
    • Asian-Australasian Journal of Animal Sciences
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    • v.27 no.10
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    • pp.1487-1492
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    • 2014
  • This research applied and evaluated a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using cytochrome b gene to detect pork contamination in meatballs from local markets in Surabaya and Yogyakarta regions, Indonesia. To confirm the effectiveness and specificity of this fragment, thirty nine DNA samples from different meatball shops were isolated and amplified, and then the PCR amplicon was digested by BseDI restriction enzyme to detect the presence of pork in meatballs. BseDI restriction enzyme was able to cleave porcine cytochrome b gene into two fragments (131 bp and 228 bp). Testing the meatballs from the local market showed that nine of twenty meatball shops in Yogyakarta region were detected to have pork contamination, but there was no pork contamination in meatball shops in Surabaya region. In conclusion, specific PCR amplification of cytochrome b gen and cleaved by BseDI restriction enzymes seems to be a powerful technique for the identification of pork presence in meatball because of its simplicity, specificity and sensitivity. Furthermore, pork contamination intended for commercial products of sausage, nugget, steak and meat burger can be checked. The procedure is also much cheaper than other methods based on PCR, immunodiffusion and other techniques that need expensive equipment.

Prevalence of High Risk Human Papillomavirus Infection with Different Cervical Cytological Features among Women Undergoing Health Examination at the National Cancer Institute, Thailand

  • Laowahutanont, Piyawat;Karalak, Anant;Wongsena, Metee;Loonprom, Komson;Pukcharoen, Phuttalak;Jamsri, Paphawin;Sangrajrang, Suleeporn
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.14
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    • pp.5879-5882
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    • 2014
  • High-risk (HR) human papillomavirus (HPV) testing is important in cervical cancer screening for triage colposcopy. The objective of the study was to evaluate the prevalence of HR HPV infection with different cervical cytological features among women undergoing health examination. A total of 2,897 women were retrospectively evaluated between May 2011 to December 2011. DNA was extracted from residual specimens collected during routine liquid-based cytology tests at the National Cancer Institute. Overall, HR HPV prevalence was 9.3% including 1.6% of HPV-16 and 0.4% of HPV-18. Of all 270 HPV positive samples, 211 (78.1% were HR-HPV non 16/18; 47 (17.4%) were HPV-16 and 12 (4.4%) were HPV-18. The prevalence of HPV infection was similar in all age groups, although a higher rate was observed in women age 31-40 years. Among women with normal cytology, HR HPV positive were found in 6.7%. In abnormal cytology, HR HPV were found 46.7% in atypical squamous cells (ASC), 54.8% in low-grade squamous intraepithelial lesions (LSIL) and 80.0% in high-grade squamous intraepithelial lesions (HSIL). HPV-16 was detected in 8.6%, 6.4% and 12.0% of ASC, LSIL and HSIL, respectively. The results of this study provide baseline information on the HPV type distribution, which may be useful for clinicians to decide who should be monitored or treated more aggressively.