Annual Report 2003

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lypopolysaccaride (LPS) molecules. As aresult, the membrane becomes permeable.EDTA and its metal salts are used as foodadditives that stabilize food colors. Ferricsodium EDTA (FeNaEDTA) is recommendedas an iron supplement for iron deficiencyanemia. The inhibitory activity of EDTAitself against H. pylori was investigated inthis study.H. pylori (ATCC 43504), the type culturestrain, was grown in Tryptic soy broth supplementedwith bovine serum at various concentrationsof disodium EDTA (Na 2 EDTA) orFeNaEDTA using a microtiter plate. After5-day incubation in anaerobic conditions at37˚C, optical density of culture was measured.Growth of H. pylori was inhibited by0.025 mM Na 2 EDTA (Fig. 3), and also by 0.1mM of FeNaEDTA. The effect of both agentswas bactericidal in contrast to Escherichiacoli and Staphylococcus aureus, whosegrowth was inhibited bacteriostatically at upFig. 3Inhibitory effect of Na 2 EDTA against growth of H. pylori.to 10 mM of Na 2 EDTA. Na 2 EDTA seemed toact on H. pylori during a logarithmic phase(after 48 hours from inoculation), where synthesisof the cell membrane which needscations such as Mg 2+ and Mn 2+ is active.Inhibition of enzymatic reactions concernedwith membrane synthesis with permeationof membrane by chelating of Mg 2+ bound toLPS may be the key reactions in this inhibition.Since the stomach volume of a humanbeing is about 1.5 L, growth of H. pylori canbe inhibited with 14 mg of Na 2 EDTA or with63 mg of FeNaEDTA. This amount ofNa 2 EDTA is much lower than the acceptabledaily intake of EDTA (2.5 mg/kg of asCaNa 2 EDTA). Estimation of the effect ofNa 2 EDTA in vivo remains to be clarified, andrequires a neutralizer because the acidicenvironment of stomachs reduces the chelatingactivity of Na 2 EDTA. However, Na 2 EDTAmight be a good choice to cure ulcers causedby H. pylori.Development and characterization of40 dinucleotide microsatellites in thedog genomeTo establish an accurate method forparentage testing in dogs, microsatelliteDNA repeat length polymorphism makerswere developed. A (CA)n-enriched library ofthe dog was constructed using the method ofTakahashi et al. (2001). The genomic DNAof the library was derived from a maleBeagle. Clones from the library containing300-700 bp inserts produced by sonication ofgenomic DNA were transformed into TG1competent cells (#200123, Stratagene, LaJolla, CA, USA). Microsatellite-containingclones were detected by hybridization todigoxigenin (DIG)-labeled (CA)12 oligonucleotideprobe using DIG nucleic acid detectionkits (Roche Diagnostics GmbH,Mannheim, Germany). Of the 384 cloneschosen from the (CA)n-enriched library, 246clones (64%) gave strongly positive signalswith the probe. A total of 192 CA repeat-positiveclones were subjected to sequencinganalysis and in 76 clones polymerase chain68 Annual Report 2003
eaction (PCR) primer pairs to detectmicrosatellite polymorphisms could be developed.Of the 76 clones, 55 PCR primer pairswere 18-25 bp long and were designed withthe assistance of Primer Premier software(Premier Biosoft International, Palo Alto,CA, USA) so as to amplify DNA fragments inthe range of 80-400 bp. Forty of the 55primer pairs were available for PCR amplificationof each locus and were examined forfurther microsatellite polymorphisms.Polymorphism: Allelic polymorphism wasdetermined by performing PCR on DNA from32 unrelated Beagle individuals. The PCRproducts were electrophoresed on an ABIPRISM TM 377 DNA Sequencer (Perkin-Elmer) and analyzed using Gene-Scan (Ver.Table 1 Indices of 40 canine microsatellite markersAverage S.D. RangeNumber of alleles 4.8 1.7 2 - 9Observed heterozygosity 0.55 0.176 0.061 - 0.791Probabilities of paternity exclusion 0.328 0.144 0.031 - 0.5873.0) and Genotyper (Ver.2.1) programs(Perkin-Elmer). The indices of variability,observed number of alleles, observed heterozygosities(Ho) and probabilities of paternityexclusion (PE) of 40 markers are shown(Table 1).The final combined PEs in Beagle was0.999999952. The results suggest that themarkers can be applied for routine parentagetesting in dogs.Annual Report 2003 69
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Message from the PresidentIn cooper
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¡X-ray crystallographic studies of
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Topics of Research in This YearComp
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Elucidation of genomic structure ar
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the total library) were mapped onto
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observed among the ribosomal protei
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molecular analyses and morphologica
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with 5 IU equine chorionic gonadotr
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serious problem in rice production
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exogenous genes into a hymenopteran
Page 23 and 24: Isolation and characterization of B
Page 25 and 26: oids could be regenerated from endo
Page 27 and 28: Quantitative trait locus analyses o
Page 29 and 30: ion, induces hypercholesterolemia,
Page 31 and 32: is an important source for amino ac
Page 33 and 34: In this study, 1.4 and 1.6 kb-long
Page 35 and 36: aqueous solution using cyanuric chl
Page 37 and 38: Developing a waste selection device
Page 39 and 40: decreased every five days by 5˚C,
Page 41 and 42: ticides. If the viral dose that kil
Page 43 and 44: In addition, the number of crown ro
Page 45 and 46: Molecular structure of the GARP fam
Page 47 and 48: X-ray crystallographic studies ofSt
Page 49 and 50: Rapid and high resolution QTL analy
Page 51 and 52: Plant regeneration system through m
Page 53 and 54: Table 1 Transformation Efficiency a
Page 55 and 56: from the tissues of mutants. The In
Page 57 and 58: the regenerated plants were transpl
Page 59 and 60: plants and fungi, and RNA interfere
Page 61 and 62: eeding, classification of microbes
Page 63 and 64: efforts are made to obtain single m
Page 65 and 66: was carried out by shotgun sequenci
Page 67 and 68: epetitive sequence specific to Oryz
Page 69 and 70: 75 µM of ABA at 25ºC. More than 8
Page 71 and 72: Change of the molecular weightforms
Page 73: A unique landrace group recognizedb
Page 77 and 78: This was the first successful case
Page 79 and 80: to disappear by degradation or dilu
Page 81 and 82: tained 1933 independent genes and b
Page 83 and 84: that antigen-specific regulatory T
Page 85 and 86: neural activity patterns evoked by
Page 87 and 88: considerable effect on the suscepti
Page 89 and 90: Insect Genetics and Evolution Depar
Page 91 and 92: activity, which was found to detoxi
Page 93 and 94: Insect Biomaterial and Technology D
Page 95 and 96: peptides obtained from sericin a re
Page 97 and 98: DNA markers for Nid-1, a resistance
Page 99 and 100: or carotenoid pigment. When sericin
Page 101 and 102: Plant Science DivisionThe Plant Sci
Page 103 and 104: mutants have been studied in relati
Page 105 and 106: have verified that candidate genes
Page 107 and 108: and C, connected by a distorted typ
Page 109 and 110: mammalian cells, respectively, inhi
Page 111 and 112: undle sheath cells (BSCs), the land
Page 113 and 114: (OsNHX1), barley (HvNHX1) and baril
Page 115 and 116: nation rates. However, in our attem
Page 117 and 118: Yeast two-hybrid assay showed inter
Page 119 and 120: and a few NADPH-cytochrome P450 oxi
Page 121 and 122: the R0(53%).Foreign genes (35S + np
Page 123 and 124: gamma ray from 44 TBq 60 Co was use
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List of PublicationOriginal Papers1
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35 Fujisaki S, Mizoguchi Y, Takahas
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71 Imai K, Khandoker MY, Yonai M, T
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107 Kiuchi S, Inage Y, Hiraiwa H, U
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147 Miyamoto Y, Sakumoto R, Sakabe
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184 Ochi A, Hossain KS, Magoshi J,
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219 Takahashi M, Nagai T, Okamura N
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258 Watanabe H, Nakashima K, Saito
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Author Department Paper.No.Yasushi
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Author Department Paper.No.Sadao Wa
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Monograph1 Magoshi J, Nakamura S (2
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on Lepidopteran Genomics”.NIAS-CO
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tion of the high-quality draft sequ
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Executive Members andResearch Staff
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Insect Growth Regulation Laboratory
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Sericultural Science LaboratoryMole
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Members of NIAS EvaluationComittee(
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Annual Report 2003 153
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Annual Report 2003

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