• Title/Summary/Keyword: RFLP.

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Investigation of Helicobacter pylori cagE Locus Diversity on Interleukin-8 Induction in AGS Cell-line (Helicobacter pylori cagE 유전자 다형성이 위상피세포주에서 Interleukin-8 유도능에 미치는 영향)

  • Lee Su-Min;Lee Hak-Sung;Lee In-Suk;Co Yu-Kyung;Han Hae-Won;Oh Jung-Hwan;Park Jae-Myung;Choi Myung-Gyu;Chung In-Sik
    • Journal of Life Science
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    • v.14 no.6 s.67
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    • pp.906-912
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    • 2004
  • Helicobacter pylori infection is highly prevalent, as high as 2/3 of whole population infected, in Korea. H. pylori infection initiates inflammation by induction of interleukin-8 through type IV secretion of CagA. It was recently suggested that induction failure of IL-8 is not associated with defect in cag PAI but associated with cagE locus diversity. This study was designed to investigate ability of 11-8 in-duction according to sequence variation within the cagE gene, cagA TP motifs and vacA m-types in vitro study using AGS cell-line, and to evaluate its association with different clinical outcome. Seventy-four H. pylori stains were isolated from 23 patients with gastric cancer (Ca), 24 subjects with gastritis (G) and 27 patients with duodenal ulcer (Du) in Kangnam St. Mary's Hospital, Seoul, Korea. cagE gene diversity was confirmed by the PCR-RFLP methods with MboI/NlaIII and tyrosine phosphate motifs (TPMs) of cagA was determined TPM-A and C by using DdeI/Tsp5091 restriction enzyme and TPM-B was determend by Real time PCR the method of Owen et al. and IL-8 was measured by ELISA assay. IL-8 activity was positively detected in 59 among 74 strains $(79.7\%)$. IL-8 secretion was significantly increased in MboI A and MboI B type compared to MboI C type and in MboI/NlaIII A-C and B-C type than C-C type. 1L-8 activity was not associated with either the number or composition of cagA tyrosine phosphorylation motifs and vacA m-type. There was no significant difference in IL-8 activity among patient groups. cagE gene diversity is thought to be mainly associated with the induction of IL-8 in H. pylori infection.

A Study of Genetic Polymonhisms of HLA-class I and II Genes Using Polymerase Chain Reaction (중합효소연쇄반응을 이용한 HLA-class I, II 유전자군의 유전적 다형성에 관한 연구)

  • Kyung-Ok Lee
    • Biomedical Science Letters
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    • v.4 no.1
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    • pp.11-25
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    • 1998
  • The HLA genes located in the short arm of chromosome 6 specify heterodimeric glycoproteins involved in the regulation of the immune response. Recently, in the elucidation of HLA polymorphism, serological and cellular typing methods have been replaced by DNA typing using polymerase chain reaction (PCR). The purpose of this study was to establish the HLA DNA typing methods and determine gene frequencies of HLA molecules in Koreans. PCR-SSP (sequence specific primers) and PCR-RFLP (restriction fragment length polymorphism) techniques were used for the analysis of HLA-A, -B, -C, DRBl genes and HLA-DQAl, DQBl, DPBl genes, respectively. The results of B-lymphoblastoid cells used for control experiment were consistent with the previous data identified in the 11th International Histocompatibility Workshop. Seventeen, 23, 16, 8, 16, 13 and 37 types of HLA-A, B, C, DQAl, DQBl, DPBl and DRBl alleles were found, respectively, in a total of unrelated 120 Korean individuals. The most frequent HLA alleles were $A^*$02 (27.0%), B$^*$40 (17.6%), Cw$^*$01 (19.2%), DQAl$^*$0301 (32.1%), DQBl$^*$0303 (12.9%), DPBl$^*$0501 (31.3%) and DRBl$^*$1501 (9.2%) among Koreans. This study shows that DNA typing method using PCR technique is a relatively simple, fast and practical tool for the determination of the HLA-class I and II genes. Moreover, the data of HLA gene frequencies could be useful for the Korean database before clinical applications, including organ and unrelated bone marrow transplantation, anthropological study, disease association and individual identification.

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Analysis of Vasopressin Receptor Type 2(AVPR2) Gene in a Pedigree with Congenital Nehrogenic Diabetes Insipidus : Identification of a Family with R202C Mutation in AVPR2 Gene (신성요붕증 가계에서 바소프레신 V2 수용체(AVPR2) 유전자 분석 : AVPR2 유전자 R202C 돌연변이의 발견)

  • Park June-Dong;Kim Ho-Sung;Kim Hee-Joo;Lee Yoon-Kyung;Kwak Young-Ho;Ha Il-Soo;Cheong Hae-Il;Choi Yong;Park Hye-Won
    • Childhood Kidney Diseases
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    • v.3 no.2
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    • pp.209-216
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    • 1999
  • Purpose : Nephrogenic diabetes insipidus (NDI) is a rare X-linked disorder associated with renal tubule resistance to arginine vasopressin (AVP). The hypothesis that the defect underlying NDI might be a dysfunctional renal AVPR2 has recently been proven by the identification of mutations in the AVPR2 gene in NDT patients. To investigate the association of mutations in th AVPR2 gene with NDI, we analyzed the AVPR2 gene located on the X chromosome. Methods : We have analyzed the AVPR2 gene in a kindred with X-linked NDI. The proband and proband's mother were analyzed by polymerase chain reaction-single strand conformational polymorphism(PCR-SSCP) and DNA sequencing of the AVPR2 gene. We also have used restriction enzyme analysis of genomic PCR product to evaluate the AVPR2 gene. Results : C to T transition at codon 202, predictive of an exchange of tryptophan 202 by cysteine(R202C) in the third extracellular domain was identified. This mutation causes a loss of Hae III site within the gene. Conclusion : We found a R202C missense mutation in the AVPR2 gene causing X-linked NDI, and now direct mutational analysis is available for carrier screening and early diagnosis.

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Current status of Brassica A genome analysis (Brassica A genome의 최근 연구 동향)

  • Choi, Su-Ryun;Kwon, Soo-Jin
    • Journal of Plant Biotechnology
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    • v.39 no.1
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    • pp.33-48
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    • 2012
  • As a scientific curiosity to understand the structure and the function of crops and experimental efforts to apply it to plant breeding, genetic maps have been constructed in various crops. Especially, in the case of Brassica crop, genetic mapping has been accelerated since genetic information of model plant $Arabidopsis$ was available. As a result, the whole $B.$ $rapa$ genome (A genome) sequencing has recently been done. The genome sequences offer opportunities to develop molecular markers for genetic analysis in $Brassica$ crops. RFLP markers are widely used as the basis for genetic map construction, but detection system is inefficiency. The technical efficiency and analysis speed of the PCR-based markers become more preferable for many form of $Brassica$ genome study. The massive sequence informative markers such as SSR, SNP and InDels are also available to increase the density of markers for high-resolution genetic analysis. The high density maps are invaluable resources for QTLs analysis, marker assisted selection (MAS), map-based cloning and comparative analysis within $Brassica$ as well as related crop species. Additionally, the advents of new technology, next-generation technique, have served as a momentum for molecular breeding. Here we summarize genetic and genomic resources and suggest their applications for the molecular breeding in $Brassica$ crop.

Morphological Characteristics and URP-PCR Analysis of Hypocrea sp., a Weed Mould of Oyster Mushroom Cultivation (느타리버섯 유해균 Hypocrea sp.의 형태 및 URP-PCR 분석)

  • Seo, Geon-Sik;Kim, Byung-Ryun;Park, Myeung-Soo;Kim, Min-Kyung;Yu, Seung-Hun
    • The Korean Journal of Mycology
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    • v.30 no.2
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    • pp.86-94
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    • 2002
  • Recently a serious outbreak of weed mould caused by a species of Hypocrea occurred in oyster mushroom (Pleurotus ostreatus) substrates in Korea. The disease was characterized by a rapid infestation of the oyster mushroom substrates by Hypocrea sp. and subsequent inhibition of fructification of the mushroom. In spite of it's serious losses to the oyster mushroom industry in Korea, etiology and ecology of the disease have not been studied. Morphological characteristics of the fungus were examined and molecular characteristics of the fungus were compared with those of the green moulds (Trichoderma spp.) isolated from oyster mushroom bed. Stromata formed superficially on suface of the substrates were pulvinate to effuse or irreguler, initially white but becoming yellowish brown, measuring $6.0{\sim}13.0{\times}3.0{\sim}11.0mm$. Perithecia were globose to subglobose, immersed in stroma, $223{\sim}263\;(Ave.239.9){\times}167.3{\sim}231\;(Ave.204.1){\mu}m$ in size. Asci were unitunicate, cylindrical, nonamyloid, $82.7{\sim}124.8\;(Ave.103.3){\times}4.1{\sim}5.1\;(Ave.4.9){\mu}m$ in size, 16 part-spored. Ascospores were bullet-shaped or somewhat oblong, hyaline, bicellular, roughened or warted, $5.4{\sim}7.4\;(Ave.6.5){\times}3.6{\sim}5.5\;(Ave.4.7){\mu}m$ in size. This fungus readily form the stroma on PDA. Mycelia on PDA nearly invisible and without cottony aerial mycelium. Optimum temperature for mycelial growth of this fungus was $25^{\circ}C$ on PDA and its growth rate was 15 mm per day. This species did not grow at below 10 and above $35^{\circ}C$. Phialides in culture enlarged in the middle and aggregated to penicillate type. They were very variable, shorted ampulliform and occasionally curved when matured, but cylinderical when young, measuring $11.9{\sim}24.3\;(Ave.\;14.7){\times}2.9{\sim}3.9\;(Ave.\;3.4){\mu}m$ when matured and $7.2{\sim}14.0\;(Ave.\;10.8){\times}2.8{\sim}4.9\;(Ave.\;3.5){\mu}m$ when young. Phialosopres were ovoid to ellipsoid, smooth, measuring $3.5{\sim}7.2\;(Ave.\;4.5){\times}2.6{\sim}3.3\;(Ave.\;2.9){\mu}m$. Nineteen isolates of Hypocrea sp. were analyzed on the basis of molecular characteristics and classified into phenotypic groups. On the basis of RAPD, URP-PCR, the fungus was confirm to monoclonal, and was classified as a different taxon from reported species of Hypocrea and Trichoderma and supposed to be a new species not previously reported in literature.

Investigation of the Relationship between Interleukin-4 Promoter Polymorphism and Severity of Patients with Bronchial Asthma (천식 환자에서 증상의 정도에 따른 IL-4 유전자 다형에 관한 연구)

  • Kang, Sea-Yong;Shim, Jae-Jeong;Cho, Jae-Yun;Kwon, Young-Hwan;Lee, Seung-Yong;Kim, Je-Hyeong;Lee, Sang-Youb;Lee, So-Ra;Han, Seon-Ae;Kim, Han-Gyum;Kang, Kyung-Ho;Yoo, Se-Hwa;In, Kwang-Ho
    • Tuberculosis and Respiratory Diseases
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    • v.45 no.3
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    • pp.529-535
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    • 1998
  • Background: Interleukin-4 plays an important role in pathogenesis of asthma, especially in developing atopy by means of switching B lymphocytes to produce IgE. It has been shown that there is polymorphism in the Interleukin-4 promoter region, transversion of cytosine to thymine at-598 from translation initiation site of IL-4 gene. There has also been quite a few works to reveal the role of the polymorphism of IL-4 gene in patients with asthma. We performed this investigation to determine the role of the polymorphism in the severity of symptoms of patients with asthma. We also examined the frequency and the type of the polymorphism in asthmatics compared with non-asthmatics as well. Method: The subjects enrolled in this study were 49 asthmatics and 33 non-asthmatics. All the asthmatics were classified as mild and moderate to severe by the NHLBI/WHO Workshop. DNA from both asthmatics and non-asthmatics was extracted, then performed ARMS(Amplification Refractory Mutation System) as well as RFLP using BsmFl restriction enzyme in order to confirm the polymorphism of Il-4 gene. Results: There was no significant difference in the occurrence of polymorphism of the IL-4 promoter sequence between asthm and non-asthma groups(P=0.7). Among those with polymorphisms, the number of C/C type was slightly more than C/T type in both asthmatics and non-asthmatics, 26 vs 21 in asthmatics and 18 vs 15 in non-asthmatics, which was, however, insignificant statistically. No significant relationship between the severity of asthma and the polymorphism was found(P=0.7). Conclusion: There was no significant difference between the severity of asthma and the IL-4 promoter polymorphism(P=0.709). Interestingly, the frequency of the polymorphism in both asthmatics as well as non-asthmatics was found to be even higher than that occurred in Caucasians. However, no significant difference in the frequency of the polymorphism was found in both groups.

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Early Matured Pear Cultivar 'Supergold' with High Quality and Greenish-white Skin for Overseas Trade (조생종 고품질 녹백색 배 '슈퍼골드')

  • Kang, Sam-Seok;Kim, Yoon-Kyeong;Cho, Kwang-Sik;Jeong, Sang-Bouk;Hwang, Hea-Seong;Kim, Myung-Su;Shin, Il-Sheob;Shin, Yong-Uk;Won, Kyeong-Ho;Choi, Jang-Jeon
    • Horticultural Science & Technology
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    • v.29 no.6
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    • pp.645-650
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    • 2011
  • Pear cultivar 'Supergold' (Pyrus pyrifolia var. culta Nakai) was originated from the cross between 'Chuwhangbae' and 'Manpungbae' with the aims of improving the fruit quality of 'Chuwhangbae' cultivar at Pear Research Station of National Institute of Horticultural & Herbal Science, Rural Development Administration in 1994. 'Supergold' was preliminarily selected in 2002 and named in 2008. The tree shows a vigorous growth habit and semi-spread characters like as 'Manpungbae'. Furthermore, it has sufficient flowers and carries abundant pollen grains, so it can also be used as a pollinator. 'Supergold' is highly resistant to black leaf spot (Alternaria kikuchiana) in the field condition. The optimum harvest time is around Sep. 11th, which is ahead of 'Whangkeumbae' about 5 days in the harvest period. The fruit shape is oblate and fruit skin color is greenish-white at harvesting time. The average weight of fruit is 570 g, and the soluble solids content is $13.6\;^{\circ}Brix$. The flesh is very soft and juicy, and renders good eating quality. Shelf life is about 6 months under the cold storage condition. To determine the self-incompatibility (SI) genotype of 'Supergold' pear cultivar, it was crossed with other cultivars of which SI genotypes have already known. The result of cross-pollinations of 'Supergold' with other cultivars showed relatively high rates of fruit set from 64.5% to 91.0%, except for the cross with pollens of 'Nijisseiki' that represented only 28.8% of fruiting rate. Although sometimes the stigma of 'Supergold' crossed with 'Hayatama', 'Chojuro', and 'Nijisseiki' showed malformed pollen tube tips, 'Supergold' is generally supposed to have cross-compatibility with all other pollen donor cultivars. It is considered that the S-allele of 'Supergold' is $S_3S_4$, which is based on the result of PCR-RFLP.

Tumor Necrosis Factor and Lymphotoxin-α Gene Polymorphism in Korean Children with Type 1 Diabetes (한국 소아 1형 당뇨병에서 종양괴사인자 및 림프독소-α 유전자 다형성)

  • Suh, Jin Soon;Park, So Young;Jung, Min Ho;Suh, Byung Kyu;Kim, Tae Gyu;Lee, Byung Churl
    • Clinical and Experimental Pediatrics
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    • v.48 no.8
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    • pp.871-876
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    • 2005
  • Purpose : Recently, it was reported that tumor necrosis factor(TNF) and $lymphotoxin-{\alpha}$($LT-{\alpha}$) gene regions might be a susceptible loci to type 1 diabetes in Japanese. The purpose of this study was to investigate the association of TNF and $LT-{\alpha}$ gene polymorphisms with disease susceptibility in Korean children with type 1 diabetes. Methods : Forty-nine Korean children with type 1 diabetes(29 girls and 20 boys) and 94 healthy Koreans were investigated in this study. Genotyping for -857T/C polymorphism in the TNF promoter region and $LT-{\alpha}$ gene polymorphism were performed by PCR-RFLP(restriction fragment length polymorphism). TNF promoter -1031C/T polymorphism was detected by allele-specific PCR. Results : The distribution of the -857T/C and -1031C/T genotype in the TNF promoter region was not different between diabetic children and the controls. The frequency of TT genotype in the distribution of TNF -1031C/T polymorphism in diabetic children with diabetic ketoacidosis(DKA) at diagnosis was significantly lower than those without DKA(P<0.05). No significant difference in the distribution of $LT-{\alpha}$ gene polymorphism was observed between diabetic children and the controls. There was no association between clinical characteristics of type 1 diabetes and $LT-{\alpha}$ gene polymorphisms. Conclusion : These results suggest that TNF promoter -857T/C and $LT-{\alpha}$ gene polymorphisms are not associated with susceptibility to type 1 diabetes in Korean children. TNF promoter -1031C/T polymorphism might be related to clinical manifestations(DKA) of type 1 diabetes.

Human Leukocyte Antigen(HLA) Genotypes and Thyroid Autoimmunity in Korean Patients with Type 1 Diabetes (한국인 제 1형 당뇨병 환자들의 HLA 유전자형 및 자가면역성 갑상선 질환의 병발 양상)

  • Kang, So Young;Shin, Chung Ho;Yang, Sei Won;Park, Myoung Hee;Yu, Jeesuk
    • Clinical and Experimental Pediatrics
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    • v.48 no.6
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    • pp.624-633
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    • 2005
  • Purpose : This study analyzed the expression of HLA-DR and DQ genotypes and anti-thyroid autoantibodies[anti-thyroid peroxidase(TPO) and anti-thyroglobulin(TG) antibodies] in Korean patients with type 1 diabetes(T1DM) to investigate the susceptible HLA alleles to T1DM in Korea and the prevalence of thyroid autoantibodies and their significance for the development of thyroid disorders. Methods : A total of 59 Korean patients with type 1 diabetes[26 males, median age 13.7 years(range 5.7-29.9 years), diabetes duration 7.6 years(-1.7-22.5 years)] were enrolled in this study, and 200 healthy Koreans without a family history of diabetes were selected as a normal control for the comparison of HLA genotypes. Seventeen patients with anti-TPO or anti-TG were followed [median duration 3.96 years(1 day-10.7 years)] with measurement of anti-TPO, anti-TG, $T_3$, $T_4$ or free $T_4$, TSH levels and physical examinations. HLA-DR and DQ genotyping were done by PCR-SSO, PCR-SSCP, PCR-RFLP and PCR-SSP methods. Results : HLA analysis showed higher frequencies of HLA-DRB1*0301, *090102 and DQB1*0201, *030302 alleles, DRB1*0301/*090102, *090102/*090102 and DQB1 *0201/*030302, *030302/*030302, *0201/ *0302 genotypes in T1DM patients compared to controls(Pc<0.05). Fifteen(25.4 percent) had anti-TPO antibody, 12(20.3 percent) had anti-TG, 17(28.8 percent) had either autoantibody and 10(16.9 percent) had both autoantibodies. No clinical or subclinical hypothyroidism developed during follow-up after the first detection of anti-thyroid autoantibody. There was no significant correlation between thyroid autoimmunity and gender, onset age of T1DM, and diabetes duration, respectively(P>0.05). Conclusion : We thought this unique HLA-DR, DQ allele distribution might be an important factor for the low incidence of T1DM in Korea. And a high prevalence of thyroid autoantibodies in these populations suggests examinations of thyroid antibodies should be performed regularly. Optimal age for the initial screening and the frequency of re-screening for associated thyroid autoimmune diseases in T1DM remains to be determined through prospective follow-up.

Apo E4 Genotype as the Alzheimer Indictor in Korean Senior Subjects Aged 50 to 64 Years Old (남녀 노인층에서 Alzheimer위험 예측 인자로써의 Apo E4 Genotype에 관한 연구)

  • Oh, Hyun-Hee;Shin, Eun-Jung;Kim, Hyun-Sook;Lim, Yun-Sook;Park, Mi-Young;Kim, Kkoch-Byul;Kim, Eun-Mee;Lee, Myoung-Sook
    • Journal of Nutrition and Health
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    • v.40 no.7
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    • pp.593-600
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    • 2007
  • Recent studies described the ${\varepsilon}4$ allele of apoE confers a two-to fourfold increased risk for late-onset Alzheimer#s disease (LOAD), but LOAD pathology does not all fit neatly around apo E. Therefore, the goal of this study was to find the association between Alzheimer and apo E4 genotype in the 107 elderly between 50 to 64 years old who visited to FHWC of Sungshin Women#s University. We conducted the questionnaire survey (general & 24 hr dietary recall), anthropometerics (BP, waist & BMI) and blood biochemistry (FBS & lipid profiles). LDL-c and HOMA-IR were calculated by Friedwald#s and Matthew#s formulas. The apo E genotyping was performed by PCR-RFLP method and subjects were divided into three allele groups (${\varepsilon}3$; wild, ${\varepsilon}2$ & ${\varepsilon}4;$ mutants). The apo E allele frequencies were 7.0% for the ${\varepsilon}2$, 83.6% for the ${\varepsilon}3$ and 9.3% for the ${\varepsilon}4$. In comparison with biochemistry characteristics by apo E genotype, FBS was significantly higher in ${\varepsilon}4(129.2{\pm}6.8)$ than that in the others (${\varepsilon}2$: $117{\pm}7.4$, ${\varepsilon}3$: $107.3{\pm}2.2)$ (p<0.01). More than forty percents of ${\varepsilon}4$ group shown the dyslipidemia [high TG (>150mg/dl) & low HDL (<40 mg/dl:male or <50 mg/dl: female)]. The cytokines levels such as IL-1 ${\beta}$, IL-6 and $TNF-{\alpha}$ were not different among three apoE alleles. After the adjusting sex, age & dietary fiber, LDL-c level was siginificantly higher in ${\varepsilon}4$ ($108.3{\pm}7.7$) than that in ${\varepsilon}2$ ($100.4{\pm}8.4$) (p<0.05). According to food intake and the recipe on the basis of 24 hr dietary recall, the elder]y with ${\varepsilon}4$ allele took higher intake frequency of the light -colored vegetable (radish, onion & cabbage) and pan-fried foods (sauteed beef and vegetables, stir-fried vienna with vegetables) than the others. We knew that the elderly with ${\varepsilon}4$ allele had been restricted the calories intakes with high dietary fiber (33.6+2.5 g/d) to maintain the normal level of FBS and LDL-c. On next study, the prevalence of Alzheimer#s disease in this population who has ${\varepsilon}4$ allele on the condition of calories restriction will be continually follow-up.