JP2020508664A - 一本鎖結合タンパク質 - Google Patents
一本鎖結合タンパク質 Download PDFInfo
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- JP2020508664A JP2020508664A JP2019545737A JP2019545737A JP2020508664A JP 2020508664 A JP2020508664 A JP 2020508664A JP 2019545737 A JP2019545737 A JP 2019545737A JP 2019545737 A JP2019545737 A JP 2019545737A JP 2020508664 A JP2020508664 A JP 2020508664A
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Abstract
Description
(i)少なくとも350mMのナトリウムイオン、
(ii)少なくとも50mMのカリウムイオン、
(iii)少なくとも150mMのマグネシウムイオン、または
(iv)少なくとも200mMのカルシウムイオン。
(i)少なくとも350mMのナトリウムイオン、
(ii)少なくとも50mMのカリウムイオン、
(iii)少なくとも150mMのマグネシウムイオン、または
(iv)少なくとも200mMのカルシウムイオン。
(i)少なくとも350mMのナトリウムイオン、
(ii)少なくとも50mMのカリウムイオン、
(iii)少なくとも150mMのマグネシウムイオン、または
(iv)少なくとも200mMのカルシウムイオン。
(i)少なくとも350mMのナトリウムイオン、
(ii)少なくとも50mMのカリウムイオン、
(iii)少なくとも150mMのマグネシウムイオン、または
(iv)少なくとも200mMのカルシウムイオン。
一本鎖DNA結合タンパク質(SSB)は、500mMのナトリウムイオンの存在下で、その最大ssDNA結合能の少なくとも50%を呈し、SSBは、配列番号1のアミノ酸配列、または配列番号1と少なくとも75%同一であるアミノ酸配列、またはその機能的断片を含む。
(i)少なくとも350mMのナトリウムイオン、
(ii)少なくとも50mMのカリウムイオン、
(iii)少なくとも150mMのマグネシウムイオン、または
(iv)少なくとも200mMのカルシウムイオン。
一本鎖DNA結合タンパク質(SSB)は、500mMのナトリウムイオンの存在下で、その最大ssDNA結合能の少なくとも50%を呈し、SSBは、配列番号1のアミノ酸配列、または配列番号1と少なくとも75%同一であるアミノ酸配列、またはその機能的断片を含む。
MARGVNKVILIGNLGDDPELRYTGSGTAVCNMSLATNETYTDSDGNEVQNTEWHDVVAWGRLGEICNEYLDKGSQVYFEGKLQTRSWEDRDNNTRYSTEVKAQEMMFLDSNRQGGADMDGFDQTRGDESLDQTRQEQPAGSSGPQPGQQASSGGEDEDTFEPDDDLPF(配列番号1)。
一本鎖DNA結合活性測定
蛍光測定
表面プラズモン共鳴
塩の混合物における結合および脱ハイブリダイズ蛍光アッセイ
SrSSB−ssDNA複合体の電子顕微鏡
オリゴマー構造
結晶化および構造決定
Claims (19)
- DNA分子を脱ハイブリダイズさせるための、500mMのナトリウムイオンの存在下で、その最大ssDNA結合能の少なくとも50%を呈する一本鎖DNA結合タンパク質(SSB)の使用であって、前記SSBは、配列番号1のアミノ酸配列、または配列番号1と少なくとも75%同一であるアミノ酸配列、またはその機能的断片を含み、前記DNA分子が、
(i)少なくとも350mMのナトリウムイオン、
(ii)少なくとも50mMのカリウムイオン、
(iii)少なくとも150mMのマグネシウムイオン、または
(iv)少なくとも200mMのカルシウムイオンのうちの1つ以上を含む溶液中に存在するかまたはその溶液に曝される、使用。 - 相補的なssDNAのハイブリダイゼーションを防ぐための、500mMのナトリウムイオンの存在下で、その最大ssDNA結合能の少なくとも50%を呈する一本鎖DNA結合タンパク質(SSB)の使用であって、前記SSBは、配列番号1のアミノ酸配列、または配列番号1と少なくとも75%同一であるアミノ酸配列、またはその機能的断片を含み、前記ssDNAは、
(i)少なくとも350mMのナトリウムイオン、
(ii)少なくとも50mMのカリウムイオン、
(iii)少なくとも150mMのマグネシウムイオン、または
(iv)少なくとも200mMのカルシウムイオンのうちの1つ以上を含む溶液中に存在するかまたはその溶液に曝される、使用。 - 前記溶液が、少なくとも500mM、好ましくは少なくとも750mMのナトリウムイオンを含む、請求項1または2に記載の使用。
- 前記溶液が、少なくとも100mM、好ましくは少なくとも300mM、好ましくは少なくとも600mMのカリウムイオンを含む、請求項1〜3のいずれか一項に記載の使用。
- 前記溶液が、少なくとも200mM、好ましくは250mMのマグネシウムイオンを含む、請求項1〜4のいずれか一項に記載の使用。
- 前記溶液が、少なくとも250mM、好ましくは300mMのカルシウムイオンを含む、請求項1〜5のいずれか一項に記載の使用。
- 前記SSBが、配列番号1のアミノ酸配列または配列番号1と少なくとも90%同一であるアミノ酸配列を含む、請求項1〜6のいずれか一項に記載の使用。
- 前記SSBが、配列番号1の17位および/または71位に置換を組み込んでいる、請求項1〜7のいずれか一項に記載の使用。
- 前記置換アミノ酸が、その側鎖にpH7.0で正電荷を担持しており、好ましくは、前記置換アミノ酸がリジンおよびアルギニンから選択される、請求項8に記載の使用。
- 前記使用が、
i)核酸増幅、精製またはシーケンシング、好ましくはナノポアシーケンシング;
ii)部位特異的突然変異誘発;
iii)試料中の核酸構造の顕微鏡検査;
iv)制限酵素消化;
v)逆転写;
vi)T4ポリメラーゼの活性の増強;または
vii)ssDNAまたはRNAのヌクレアーゼ消化からの保護の方法の間に行われる、請求項1〜9のいずれか一項に記載の使用。 - 500mMのナトリウムイオンの存在下で、その最大ssDNA結合能の少なくとも50%を呈し、かつ配列番号1のアミノ酸配列、または配列番号1と少なくとも75%同一であるアミノ酸配列、またはその機能的断片を含み、かつ17位および/または71位の前記アミノ酸が置換されている、一本鎖DNA結合タンパク質(SSB)。
- 配列番号2のアミノ酸配列を含む、請求項18に記載のSSB。
- 請求項11または12に記載のSSBをコードする、核酸分子。
- 請求項13に記載の核酸分子を含む、発現ベクター。
- 請求項14に記載の発現ベクターを含む、宿主細胞またはウイルス。
- 500mMのナトリウムイオンの存在下で、その最大ssDNA結合能の少なくとも50%を呈するSSBをコードする、配列番号4の核酸配列またはその断片を含む、核酸分子。
- 500mMのナトリウムイオンおよび緩衝液および任意により安定化剤の存在下で、その最大ssDNA結合能の少なくとも50%を呈する単離されたSSBを含む組成物であって、前記SSBは、配列番号1のアミノ酸配列、または配列番号1と少なくとも75%同一であるアミノ酸配列、またはその機能的断片を含む、組成物。
- 請求項17に記載の単離されたSSB、または請求項11もしくは12に記載の単離されたSSB、または請求項17に記載の組成物を、任意によりポリメラーゼ、洗浄緩衝液または他の緩衝液、ヌクレオチド、核酸プライマー、リガーゼ、トポイソメラーゼまたはジャイレースのうちの1つ以上と共に含む、キット。
- シーケンシング装置での使用に好適であるカートリッジ内に収容されている、請求項17に記載のSSB、または請求項11もしくは12に記載のSSB、または請求項17に記載の組成物。
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GB201512617D0 (en) | 2015-07-17 | 2015-08-26 | Francis Crick Inst The Ltd | Single-stranded DNA sensor |
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2017
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2018
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JP7273717B2 (ja) | 2023-05-15 |
US11447812B2 (en) | 2022-09-20 |
CN110291208A (zh) | 2019-09-27 |
US20230092081A1 (en) | 2023-03-23 |
KR20190120271A (ko) | 2019-10-23 |
EP3585907A1 (en) | 2020-01-01 |
WO2018154083A1 (en) | 2018-08-30 |
GB201703049D0 (en) | 2017-04-12 |
KR102636161B1 (ko) | 2024-02-14 |
CA3052727A1 (en) | 2018-08-30 |
US20190376122A1 (en) | 2019-12-12 |
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