• Title/Summary/Keyword: protein sequencing

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Human Parechovirus as an Important Cause of Central Nervous System Infection in Childhood (소아청소년기 중추신경 감염의 주요 원인으로서 Human Parechovirus의 의의)

  • Jung, Hyun Joo;Choi, Eun Hwa;Lee, Hoan Jong
    • Pediatric Infection and Vaccine
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    • v.23 no.3
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    • pp.165-171
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    • 2016
  • Purpose: Human parechovirus (HPeV) is an increasingly recognized pathogenic cause of central nervous system (CNS) infection in neonates. However, HPeV infections have not been studied in older children. This study determined the prevalence and clinical features of HPeV CNS infection in children in Korea. Methods: Reverse transcription polymerase chain reaction assays were performed using HPeV-specific, 5' untranslated, region-targeted primers to detect HPeV in cerebrospinal fluid (CSF) samples from children presenting with fever or neurologic symptoms from January 1, 2013, to July 31, 2014. HPeV genotyping was performed by sequencing the viral protein 3/1 region. Clinical and laboratory data were retrospectively abstracted from medical records and compared with those of enterovirus (EV)-positive patients from the same period. Results: Of 102 CSF samples, six (5.9%) were positive for HPeV; two of 21 EV-positive samples were co-infected with HPeV. All samples were genotype HPeV3. Two HPeV-positive patients were <3 months of age and four others were over 1 year old. While HPeV-positive infants under 1 year of age presented with sepsis-like illness without definite neurologic abnormalities, HPeV-positive children over 1 year of age presented with fever and neurologic symptoms such as seizures, loss of consciousness, and gait disturbance. The CSF findings of HPeV-positive patients were mostly within the normal range, whereas most (73.7%) EV-positive patients had pleocytosis. Conclusions: Although HPeV is typically associated with disease in young infants, the results of this study suggest that HPeV is an emerging pathogen of CNS infection with neurologic symptoms in older childhood.

Identification and Characterization of Lactobacillus salivarius subsp. salivarius from Korean Feces

  • Bae, Hyoung-Churl
    • Proceedings of the Korean Society for Food Science of Animal Resources Conference
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    • 2004.05a
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    • pp.89-119
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    • 2004
  • This study was conducted to isolate lactobacilli having probiotic characteristics to be used as health adjuncts with fermented milk products. Acid tolerant strains were selected in Lactobacilli MRS broth adjusted to pH 4.0 from 80 healthy persons (infants, children and adults). And bile tolerant strains were examined in Lactobacilli MRS broth in which 1.0% bile salt was added. By estimation above characteristics, the strains No. 27, which was isolated from adult feces, was selected and identified as Lactobacillus salivarius subsp. salivarius based on carbohydrate fermentation and 16S rDNA sequencing. It was used as a probiotic strain in fermented milk products. The pH of fermented milk decreased from pH 6.7 to 5.0 and titratable acidity increased from 0.3% to 1.0% by L. salivarius subsp. salivarius (isolation strain 20, 35, and 37), when incubated for 36 h at $37^{\circ}C$. The number of viable cell counts of fermented milk was maximized at this incubation condition. The SDS-PAGE evidenced no significant change of casein but distinct changes of whey protein were observed by isolated L. salivarius subsp. salivarius for titratable acidity being incubated by $0.9{\sim}1.0%$ at $37^{\circ}C$. All of the strains produced 83.43 to 131.96 mM of lactic acid and 5.39 to 26.85 mM of isobutyric acid in fermented products. The in vitro culture experiment was performed to evaluate ability to reduce cholesterol levels and antimicrobial activity in the growth medium. The selected L. salivarius subsp. salivarius reduced $23{\sim}38%$ of cholesterol content in lactobacilli MRS broth during bacterial growth for 24 hours at $37^{\circ}C$. All of the isolated L. salivarius subsp. salivarius had an excellent antibacterial activity with $15{\sim}25$ mm of inhibition zone to E. coli KCTC1039, S. enteritidis KCCM3313, S. typhimurium M-15, and S. typhimurium KCCM40253 when its pH had not been adjusted. Also, all of the isolated L. salivarius subsp. salivarius had partial inhibition zone to E. coli KCTC1039, E. coli KCTC0115 and S. enteritidis KCCM3313 when it had been adjusted to pH 5.7. The selected strains were determined to have resistances of twelve antibiotic. Strains 27 and 35 among the L. salivarius subsp. salivarius showed the highest resistance to the antibiotics. Purified ${\alpha}$-galactosidase was obtained by DEAE-Sephadex A-50 ion exchange chromatography, Mono-Q ion exchange chromatography and HPLC column chromatography from L. salivarius subsp. salivarius 27. The specific activity of the purified enzyme was 8,994 units/mg protein, representing an 17.09 folds purification of the original cell crude extract. The molecular weight of enzyme was identified about 53,000 dalton by 12% SDS-PAGE. Optimal temperature and pH for activity of this enzyme were $40^{\circ}C$ and 7.0 respectively. The enzyme was found to be stable between 25 and $50^{\circ}C$. ${\alpha}$-galactosidase activity was lost rapidly below pH 5.0 and above pH 9.0. This enzyme was liberated galactose from melibiose, raffinose, and stachyose, and also the hydrolysis rate of substrate was compound by HPLC. These results indicated that some of the L. salivarius subsp. salivarius (strain 27 and 35) are considered as effective probiotic strains with a potential for industrial applications, but the further study is needed to establish their use as probiotics in vivo.

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Manufacture of Cutting-Gouda Cheese using Lactobacillus rhamnosus_p1 and the Physicochemical Properties of Gouda Cheese during Ripening Periods (Lactobacillus rhamnosus_p1을 이용한 절단형 고다치즈 제조방법 및 숙성 중 품질특성)

  • Park, Jong-Hyuk;Jung, Hoo-Kil;Moon, Hye-Jung;Oh, Jeon-Hui;Lee, Joo-Hee;Kim, Myung-Kon;Na, Sang-Eon;Kim, Youn-Jeong;Hwang, Young-Tae
    • Journal of Dairy Science and Biotechnology
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    • v.32 no.2
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    • pp.131-139
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    • 2014
  • The aim of this study was to manufacture Cutting-Gouda cheese and to investigate the change in physicochemical properties of Cutting-Gouda cheese made with Lactobacillus rhamnosus_p1. Lactic acid bacteria were isolated from Gouda cheese ripened for more than 1 year. They were identified as 2 strains of L. rhamnosus, Lactobacillus casei, Lactobacillus curvatus, and Staphylococcus saprophyticus by 16S rDNA sequencing and named L. rhamnosus_p1, L. casei_p2, L. curvatus_p3, L. rhamnosus_p4 and S. saprophyticus_p5. The proteolytic activities of isolated strains against casein were measured using prepared skim milk agar plates. L. rhamnosus_p1 showed the highest proteolytic activity. Cutting-Gouda cheese was made with L. rhamnosus_p1, and its physicochemical properties (moisture, protein, fat, ash and free amino acid content) were measured during ripening periods. Because of the modified atmosphere packaging ($N_2{^-}$), there was no change in moisture, protein, fat, and ash in the experimental group. The total amount of free amino acids in the control and experimental group gradually increased during ripening periods. The sensory evaluation showed that the experimental group was preferable to the control group. This result suggests that L. rhamnosus_p1 has potential to be developed as a new starter for Gouda cheese.

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A prognosis discovering lethal-related genes in plants for target identification and inhibitor design (식물 치사관련 유전자를 이용하는 신규 제초제 작용점 탐색 및 조절물질 개발동향)

  • Hwang, I.T.;Lee, D.H.;Choi, J.S.;Kim, T.J.;Kim, B.T.;Park, Y.S.;Cho, K.Y.
    • The Korean Journal of Pesticide Science
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    • v.5 no.3
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    • pp.1-11
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    • 2001
  • New technologies will have a large impact on the discovery of new herbicide site of action. Genomics, combinatorial chemistry, and bioinformatics help take advantage of serendipity through tile sequencing of huge numbers of genes or the synthesis of large numbers of chemical compounds. There are approximately $10^{30}\;to\;10^{50}$ possible molecules in molecular space of which only a fraction have been synthesized. Combining this potential with having access to 50,000 plant genes in the future elevates tile probability of discovering flew herbicidal site of actions. If 0.1, 1.0 or 10% of total genes in a typical plant are valid for herbicide target, a plant with 50,000 genes would provide about 50, 500, and 5,000 targets, respectively. However, only 11 herbicide targets have been identified and commercialized. The successful design of novel herbicides depends on careful consideration of a number of factors including target enzyme selections and validations, inhibitor designs, and the metabolic fates. Biochemical information can be used to identify enzymes which produce lethal phenotypes. The identification of a lethal target site is an important step to this approach. An examination of the characteristics of known targets provides of crucial insight as to the definition of a lethal target. Recently, antisense RNA suppression of an enzyme translation has been used to determine the genes required for toxicity and offers a strategy for identifying lethal target sites. After the identification of a lethal target, detailed knowledge such as the enzyme kinetics and the protein structure may be used to design potent inhibitors. Various types of inhibitors may be designed for a given enzyme. Strategies for the selection of new enzyme targets giving the desired physiological response upon partial inhibition include identification of chemical leads, lethal mutants and the use of antisense technology. Enzyme inhibitors having agrochemical utility can be categorized into six major groups: ground-state analogues, group specific reagents, affinity labels, suicide substrates, reaction intermediate analogues, and extraneous site inhibitors. In this review, examples of each category, and their advantages and disadvantages, will be discussed. The target identification and construction of a potent inhibitor, in itself, may not lead to develop an effective herbicide. The desired in vivo activity, uptake and translocation, and metabolism of the inhibitor should be studied in detail to assess the full potential of the target. Strategies for delivery of the compound to the target enzyme and avoidance of premature detoxification may include a proherbicidal approach, especially when inhibitors are highly charged or when selective detoxification or activation can be exploited. Utilization of differences in detoxification or activation between weeds and crops may lead to enhance selectivity. Without a full appreciation of each of these facets of herbicide design, the chances for success with the target or enzyme-driven approach are reduced.

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Association Between the Polymorphism in the Promoter Region of Porcine A-FABP Gene and Growth Traits in Duroc Pigs (돼지 Duroc 품종에서 A-FABP promoter의 다형성과 성장형질의 연관성)

  • Han, Sang-Hyeon;Jo, In-Cheol;Lee, Jong-Eon;Kim, Hyo-Seon;Lee, Jeong-Gyu;Jeon, Jin-Tae;O, Mun-Yu;Go, Mun-Seok
    • Journal of Animal Science and Technology
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    • v.48 no.1
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    • pp.21-28
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    • 2006
  • A polymorphism was found in the promoter region of porcine adipocyte fatty acid binding protein gene(A-FABP) gene which plays a key role in the binding and transportation of free fatty acid in adipocyte and deposition of intramuscular fat. Mutation was detected a substitution(T406C) using SSCP analysis and subsequently confirmed by sequencing the fragment in Duroc pigs. This T-406C mutation might change the binding activity for transcription factor nuclear factor 1(NF1). In this population, this mutation was genotyped using HinfⅠRFLP, and found three kinds of genotypes(TT, TC, and CC) showing their frequencies of 42.3, 44.3, and 13.4%, respectively. We statistically analyzed the association between the A-FABP genotypes and growth traits and found that the body weights of the pigs containing 406C/(TC or CC) were heavier for the body weight at the age of 20 weeks than those containing genotype TT(P<0.05), but not for those at the age of 0, 3, and 10 weeks. Pigs containing genotype CC had also a higher value for the average daily gain and lower values for the date for 90kg of body weight and food conversion ratio than those of 406T/- genotype. In addition, without the significant difference of back fat thickness, there was a significant association between the existence of allele CC and lean meat and eye muscle area(P<0.05). As a result of this study, we suggest that the allele T406C in the promoter region of A-FABP gene play an important role in deposition of intramuscular fat and weight in the later growth period. This polymorphism will be an useful molecular marker for breeding of Duroc pigs.

Anti-inflammatory Activity of Antimicrobial Peptide Papiliocin 3 Derived from the Swallowtail Butterfly, Papilio xuthus (호랑나비 유래 항균 펩타이드 파필리오신 3의 항염증 활성)

  • Shin, Yong Pyo;Lee, Joon Ha;Kim, In-Woo;Seo, Minchul;Kim, Mi-Ae;Lee, Hwa Jeong;Baek, Minhee;Kim, Seong Hyun;Hwang, Jae Sam
    • Journal of Life Science
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    • v.30 no.10
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    • pp.886-895
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    • 2020
  • The development of novel peptide antibiotics with potent antimicrobial activity and anti-inflammatory activity is urgently needed. In a previous work, we performed an in-silico analysis of the Papilio xuthus transcriptome to identify putative antimicrobial peptides and identified several candidates. In this study, we investigated the antibacterial and anti-inflammatory activities of papiliocin 3, which was selected bioinformatically based on its physicochemical properties against bacteria and mouse macrophage Raw264.7 cells. Papiliocin 3 showed antibacterial activities against E. coli and S. aureus without inducing hemolysis and decreased the nitric oxide production of the lipopolysaccharide-induced Raw264.7 cells. Moreover, ELISA and Western blot analysis revealed that papiliocin 3 reduced the expression levels of pro-inflammatory enzymes, such as inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and prostaglandin E2 (PGE2). In addition, we examined whether papiliocin 3 could inhibit the expression of pro-inflammatory cytokines (interleukin-6 and interleukin-1β) in LPS-induced Raw264.7 cells. We found that papiliocin 3 markedly reduced the expression level of cytokines through the regulation of mitogen-activated protein kinases (MAPK) and nuclear factor kappa B (NF-κB) signaling. We also confirmed that papiliocin 3 binds to bacterial cell membranes via a specific interaction with lipopolysaccharides. Collectively, these findings suggest that papiliocin 3 could be a promising molecule for development as a novel peptide antibiotic.

Susceptibility Loci Associations with Prostate Cancer Risk in Northern Chinese Men

  • Wang, Na-Na;Xu, Yong;Yang, Kuo;Wei, Dong;Zhang, Yao-Guang;Liu, Ming;Shi, Xiao-Hong;Liang, Si-Ying;Sun, Liang;Zhu, Xiao-Quan;Yang, Yi-Ge;Tang, Lei;Zhao, Cheng-Xiao;Wang, Xin;Chen, Xin;Hui, Juan;Zhang, Yu-Hong;Zhu, Ling;Yang, Fan;Zhang, Yu-Rong;Yang, Ze;Wang, Jian-Ye
    • Asian Pacific Journal of Cancer Prevention
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    • v.14 no.5
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    • pp.3075-3078
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    • 2013
  • Background: KLK3 gene products, like human prostate-specific antigen (PSA), are important biomarkers in the clinical diagnosis of prostate cancer (PCa). G protein-coupled receptor RFX6, C2orf43 and FOXP4 signaling plays important roles in the development of PCa. However, associations of these genes with PCa in northern Chinese men remain to be detailed. This study aimed to investigate their impact on occurrence and level of malignancy. Methods: All subjects were from Beijing and Tianjin, including 266 cases with prostate cancer and 288 normal individuals as controls. We evaluated associations between clinical covariates (age at diagnosis, prostate specific antigen, Gleason score, tumor stage and aggressive) and 6 candidate PCa risk loci, genotyped by PCR- high resolution melting curve and sequencing methods. Results: Case-control analysis of allelic frequency of PCa associated with PCa showed that one of the 6 candidate risk loci, rs339331 in the RFX6 gene, was associated with reduced risk of prostate cancer (odds ratio (OR) = 0.73, 95% confidence interval (CI) =0.57-0.94, P = 0.013) in northern Chinese men. In addition, subjects with CX (CC+TC) genotypes had a decreased risk for prostrate cancer compared to those carrying the TT homozygote (OR =0.64, 95% CI = 0.45- 0.90, P = 0.008). The TT genotype of 13q22 (rs9600079, T) was associated with tumor stage (P=0.044, OR=2.34, 95% CI=0.94-5.87). Other SNPs were not significantly associated with clinical covariates in prostate cancer (P > 0.05). Conclusions. rs339331 in the RFX6 gene may be associated with prostate cancer as a susceptibility locus in northern Chinese men.

A case of Hyper-IgE syndrome with a mutation of the STAT3 gene (STAT3유전자 돌연변이 검사로 확진된 고면역글로불린E 증후군 1례)

  • Kang, Ji-Man;Suh, Jung-Min;Kim, Ji-Hyun;Kim, Hee-Jin;Kim, Yae-Jean;Lee, Hun-Seok;Shin, Young-Kee;Ahn, Kang-Mo;Lee, Sang-Il
    • Clinical and Experimental Pediatrics
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    • v.53 no.4
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    • pp.592-597
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    • 2010
  • Hyperimmunoglobulin E syndrome (HIES) is a rare immunodeficiency disease which is characterized by high serum IgE levels, eczema, and recurrent infections. Herein we present the case of a patient with HIES associated with STAT3 gene ($stat3$) mutation. A 16 year-old girl was admitted to our hospital due to hemoptysis caused by pneumonia with bronchiectasis. She had a history of recurrent skin and respiratory tract infections, such as pneumonia caused by MRSA (methicillin-resistant $Staphylococcus$ $aureus$) and $Pseudomonas$ $aeruginosa$. On physical examination, a broad round shaped nose, oral thrush, and chronic eczematous skin rash over her whole body were found. Laboratory data showed an elevated eosinophil count ($750/{\mu}L$) and total IgE level (5,001 U/mL). The patient's National Institutes of Health (NIH) score for HIES was 44. Direct sequencing of the STAT3 gene revealed that the patient was heterozygous for a missense mutation in the DNA binding domain of the STAT3 protein (c.1144C>T, p. Arg382Trp). HIES should be suspected in patients with recurrent infections and can be confirmed by clinical scoring and genetic analysis.

Detection of Lily symptomless virus, Lily mottle virus, and Cucumber mosaic virus from Lilium Grown in Korea by RT-PCR (RT-PCR법을 이용한 백합 바이러스 LSV, LMoV, CMV의 검출)

  • Lim, Ji-Hyun;Bae, Eun-Hye;Lee, Yong-Jin;Park, Sung-Han;Lee, Kyu-Jun;Kim, Sae-Ro-Mi;Jung, Yong-Tae
    • Korean Journal of Microbiology
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    • v.45 no.3
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    • pp.251-256
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    • 2009
  • Leaf samples and bulbs showing characteristic symptoms of virus infection were collected from Gang-won, Chung-nam, and Jeju Province of Korea in 2008-2009. Three viruses, Lily symptomless virus (LSV), Lily mottle virus (LMoV), and Cucumber mosaic virus (CMV) were detected by RT-PCR. Virus-infected plant samples were identified; 12 plants with LSV, 20 plants with LMoV, and 1 plant with CMV. Of the twelve LSV infected samples, seven samples were found to be mix-infected with LMoV and LSV. Symptoms of LMoV and LSV mixed infection were fairly severe, like as vein clearing, leaf curling, leaf mottling, leaf mosaic, and yellow streaking. Mixed infection with LMoV and LSV was also found in lily bulbs which have been stored under unfavorable environmental conditions. LMoV predominated in our tests, whereas spread of Lilyvirus X (LVX) was not found. The nucleotide sequences of coat protein (CP) region of seven isolates (4 LMoV, 2 LSV, and 1 CMV) were compared with the corresponding regions of LMoV (AJ564636), LSV (AJ516059) and CMV(AJ296154). The nucleotide sequence homologies between reference viruses and seven isolates were 95-99%. Complete sequencing of seven isolates is necessary to obtain more information on the molecular characteristics of these viruses as well as to increase sensitivity and rapidity of viral detection.

Polymorphisms of TAS1R3 and GNAT3 Genes Are Associated with Patients with Taste Disorder (미각장애와 TAS1R3 및 GNAT3 유전자의 다형성과의 연관성)

  • Bae, Jae-Woong;Kim, Un-Kyung;Kwon, Tae-Jun;Choi, Su-Jin;Ye, Mi-Kyung
    • Journal of Life Science
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    • v.21 no.3
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    • pp.412-416
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    • 2011
  • Taste sensation plays a crucial role in selecting and ingesting foods with different qualities which convey information about their nutrient content and/or safety. Sweetness is one of the five modalities in humans and serves as an energy resource for metabolism. There are reports on allelic polymorphisms which influence perception of sweetness in mice and humans. Since the influence of genetic factors on taste disorder has not been studied, we investigated the association of genetic polymorphisms in TAS1R3 and guanine nucleotide binding protein, alpha transducing 3 (GNAT3) genes and taste disorder. A total of 150 individuals composed of 50 patients with taste disorder and 100 healthy controls were recruited for the study and PCR-mediated directing sequencing method was used to genotype for two different single nucleotide polymorphisms (SNPs) - rs307355 (T>C) and rs35744813 (T>C) in the TAS1R3 gene, and rs7792845 (T>C) and rs1524600 (C>T) in the the GNAT3 gene. The allele and genotype frequencies of rs307355 and rs35744813 in the TAS1R3 gene showed a significant association between patients with taste disorder (p=0.022 and p=0.013 in both of SNPs, respectively). In addition, the frequency of T-T haplotype in the TAS1R3 gene was higher in taste disorder cases than in the controls (OR, 1.93: 95%. CI, 1.09-3.39, p=0.022). In the GNAT3, the genotype frequency of rs7792845 in the patients was also different from the controls (p=0.048), but allele frequency was not significantly associated in either group. Our result provides the frequencies of SNPs and haplotypes of the TAS1R3 and GNAT3 genes for the fundamental information of nutrigenetics in perception of the taste of sweetness in the Korean population. Also, the study suggests that the allelic polymorphisms of TAS1R3 and GNAT3 genes may be useful as a molecular marker for evaluating patients with taste disorder. Further studies with large samples are required to clarify our observation.