Table 1 Detection bacterial strains used in this study *1; Methicillin resistant strains(mrsa)
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- ゆきひら よしなが
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1 Key words: PCR, ribosomal RNA, bacterial contamination
2 Table 1 Detection bacterial strains used in this study *1; Methicillin resistant strains(mrsa)
3 PCR法 Fig. 1 Agarose gel ribosomal DNA Amplification primers (rdna) from using ethidium s are lar markers as resistant Typhi, 4; 6; Verotoxin (VTEC), 7; (EIEC), 10: C. in A, producing 3; each 5; mixed, E. coli 8; E. rdna coli A. Mlul, Eco molecular 所 の み で 切 断 さ れ,MluIで 541と347bp,EcoRIで 3; RI 1% A (E). on to E indi- type-1 (C), Lanes 0; individual to agarose parahaemolyticus MRSA HindlIl, 37 Ž. used dysenteriae performed marker Panels PCR- at strains, respectively. s (0./HindIII P. *, ) は は732と156bp,HindIIIで は758と130bpの る.そ PCR 1 to 1ƒÊl 60min TenƒÊl mixture fragments (B), (D) s water for metnod. sobria follows. 5 `10units) PCR- 類 の 制 限 酵 素 で 処 理 す る と,い HindIII. concentrated distilled 10ƒÊl or ten-fold incubated gel- as (about incubation, (A), ず れ の 場 合 も1ヵ 16ƒÊl aeruginosa で は 使 用 し た3種 3ƒÊl buffer, cates digested EcoRI performed product, confirm jciuni. target separately Mlul, endonuclease After endonuclease 2; endonuclease. PCR fragments amplified gel (888bp) Enteroinvasive Agarose restriction each (MRSA), parahaemolyticus, 9; An molecu- ), Paratyphi 2 restriction 11; 497 rdna agarose-gel 1 aureus cholerae, Typhimurium, 1% samples : s (ƒé/hindiii methicillin Fig. observed follows PCR- detection microorganisms. staining bromide. for に よ る細 菌汚 染 確 認 小 断片 とな る こ とがわ か って い こで そ れ らの 制 限 酵 素 の 認識 部 位 の有 無 に よ る 群 別 で は,全 て の 酵 素 に 感 受 性 のEcol4 Shigella spp.のi群,上 記2菌 EcoRIに 感 受 性 の あ るSalmonellaを るEnterobacteriaceae科 のII群,Mlulの 種 以 外 のMluI, は じめ とす お よ びAeromonas み で 切 断 さ れ るVibrio sppspp.とp. damselaのiii群,p.aeruginosa,c.jeiuni,c.fetus で み ら れ る よ う な3種 類 さ れ た.な 類 に 感 受 性 が な いIV群 おC.fetusで 合 に は,切 断 前rDNAサ はMlulで グ ラ ム 陽 性 菌 のMRSAで は み ら れ な か っ た. は グ ラ ム陰 性 菌 と同 じ 増 幅 さ れ た が,MluIに 受 性 が な く,EcoRI,HindIIIに I IV群 切 断 した 場 片 中 に は,HindIIIに2ヵ 認 識 部 位 を 持 つ こ と を 意 味 す る3断 平 成6年4月20日 は感 は 感 受 性 が あ り, と は 異 な っ た パ タ ー ン(V群)を ま た こ のrDNA断 I群 のE.coliやShigella 示 し た. 所 の 片 が 確 認 され spp-の 塩 基 配 列 とは 違 っ た も の で あ っ た(Fig.2). イ ズ よ り小 さ く な っ た 泳 動 像 が み ら れ た がC.jejuniで サ イ ズ の 標 的DNAが に分 考 察 細 菌 の タ ン パ ク 合 成 に 関 与 す る リボ ソ ー ム は, 近年 系統 発 生的 研 究 のた め そ の構造 解 析 が行 わ れ1),そ れ を 構 成 し て い るrRNAは,DNAに 比べ て 細 菌 細 胞 内 に 大 量 に 存 在 し て い る こ と か ら, PCRを 行 う に はDNAを 対 象 とす る よ り も 効 率 良 く実 施 で き る.rRNAに の3種 が 知 ら れ て い る が,そ は5S,16Sと23SRNA の 大 き さ か ら16Sが
4 Table 2 Restriction endonuclease patterns groups bacteria examined Verotoxin-producing E. coli; 2, Enteroinvasive E. coli; 1 Enteropathogenic E. coli; 1 strains examined. dysenteriae 1; flexneri 3a; 1 sonnei; 2 strains examined. Typhi; 2, Paratyphi A; 2, Enteritidis; 1 Typhimurium; 2 strains examined. C. freundii; 2 C. diversus; 5 strains examined. P. mirabilis; 1 P. vulgaris; 1 strains examined. K. pneumoniae; 2 K. oxytoca; 1 strains examined. A. hydrophila; 2 A. sobria; 1 strains examined. cholerae O1, strain 569B included. most clear DNA fragment this enzyme recognized a small fragment. This result not observed in case C. jejuni strains. two DNA fragments Hind III endonuclease different from its group I.
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