Chiba/Flagella/Making FliC-his

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< Chiba
Revision as of 03:20, 27 October 2007 by Risa (Talk | contribs)
  1. pUC19 vectorにFliCをコードする遺伝子を挿入→pUC19-FliC(島田さんから)
  2. エキサイトPCR
  3. design insert(ヒスチジンタグ)
  4. pUC19-FliC vector とinsertオリゴをライゲーション.
  5. GelCheck

[混ぜ表]

  • Template1μL
  • Primer(FとR)5μL
  • 10×buffer10μL
  • dNT
  • design primer
    • [PCR表]FliCの可変領域のうち、フィラメントの外側部分から５か所を選び、プライマーを設計(207,228,240,252,267)
      • pUC19-FliCをpUC19-FliC-vectorに
  • six histidine loop peptide ("His-loop")
  • アミノ酸配列-Ser-Ser-His-His-His-His-His-His-Ser-Ser
  • encoded by the oligo 5'-TCT-TCT-CAT-CAT-CAT-CAT-CAT-CAT-TCC-TCC-3'
  • and anti oligo 5'-GGA-GGA-ATG-ATG-ATG-ATG-ATG-ATG-AGA-AGA-3'
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