HRP20201452T1 - Izoliranje nukleinskih kiselina - Google Patents

Izoliranje nukleinskih kiselina Download PDF

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HRP20201452T1
HRP20201452T1 HRP20201452TT HRP20201452T HRP20201452T1 HR P20201452 T1 HRP20201452 T1 HR P20201452T1 HR P20201452T T HRP20201452T T HR P20201452TT HR P20201452 T HRP20201452 T HR P20201452T HR P20201452 T1 HRP20201452 T1 HR P20201452T1
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concentration
volume
ethanol
solid support
water
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HRP20201452TT
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Stephen John Minter
Georgios Patsos
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RevoluGen Limited
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    • C12N15/00Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
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    • C12N15/00Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09Recombinant DNA-technology
    • C12N15/10Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/1003Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
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    • C12Q1/00Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6806Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay
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    • C12Q2527/00Reactions demanding special reaction conditions
    • C12Q2527/137Concentration of a component of medium

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Claims (11)

1. Postupak izoliranja nukleinskih kiselina koje sadrže DNK iz biološkog materijala, postupak koji obuhvata korake: (i) vršenja liziranja biološkog materijala vodenim rastvorom koji sadrži litij u koncentraciji od 0.05 do manje od 1M, kelatni agens, i surfaktant za proizvodnju vodene lizirane kompozicije, (ii) tretiranja lizirane kompozicije čvrstim nosačem u prisustvu otopljenog kaotropnog agensa u koncentraciji od 0.05 do 2M i 25% do 60% po zapremini u tekućini otopljenog C1-3 alkanola, pri čemu je navedeni čvrst nosač sposoban za imobilizaciju DNK, (iii) odvajanja čvrstog nosača od tekućine, (iv) tretiranja čvrstog nosača prvom otopinom za ispiranje koji sadrži litij u koncentraciji od 0.05 do 1M, pri čemu navedena prva otopina za ispiranje sadrži 20% do 90% po zapremini otopljenog C1-3 alkanola, (v) odvajanja čvrstog nosača od prve tekućine za ispiranje, (vi) tretiranja čvrstog nosača drugom tekućinom za ispiranje koja sadrži najmanje 80% po zapremini C1-3 alkanola, pri čemu je ravnoteža ako je prisutna voda, (vii) odvajanja čvrstog nosača od druge otopine za ispiranje, i (viii) eluiranja DNK sa čvrstog nosača.
2. Postupak prema patentnom zahtjevu 1 gdje vodena otopina opisana u koraku (i) sadrži litij u koncentraciji od 0.05 do 0.9M.
3. Postupak prema patentnom zahtjevu 1 gdje se otopina za lizu suštinski sastoji od vode, litij klorida u koncentraciji od 0.7 do 0.9M, EDTA ili natrijeve soli istog u koncentraciji od 5 do 15mM i 0.9 do 1.1% po masi SDS, opcionalno gdje u koraku (ii) kaotropni agens je gvanidinij klorid u koncentraciji od 0.6 do 0.7M i alkanol je etanol prisutan u količini od 35-40% po zapremini tekućine.
4. Postupak prema patentnom zahtjevu 1 gdje se otopina za lizu sastoji suštinski od vode, litij klorida u koncentraciji od 0.2 do 0.3M, EDTA ili natrijeve soli istog u koncentraciji od 1 do 5mM, gvanidinij klorida u koncentraciji od 0.6 do 0.7M, SDS u količini od 0.2 do 0.4% po masi i 35-40% po zapremini etanola.
5. Postupak prema bilo kojem od patentnih zahtjeva 1 do 4 gdje se prva tekućina za ispiranje suštinski sastoji od a) vode, litij klorida u koncentraciji od 0.3 do 0.6M, i 45% do 55% po zapremini etanola; ili b) vode, litij klorida u koncentraciji od 0.6 do 1M, i 45% do 55% po zapremini etanola.
6. Postupak prema bilo kojem od patentnih zahtjeva 1 do 5 gdje a) druga tekućina za ispiranje se suštinski sastoji od 80% do 95% po zapremini etanola i vode i/ili b) čvrsti nosač sadrži silicij-dioksid; i/ili c) korak (iii) se vrši u epruveti za centrifugu u kojoj navedeni čvrst nosač je filter i koraci (iii), (v), (vii) i (viii) se vrše uz centrifugiranje.
7. Postupak prema patentnom zahtjevu 6 (c) gdje filter sadrži vlakna borosilikatnog stakla, opcionalno gdje je veliki broj navedenih filtera osiguran u epruveti za centrifugu, opcionalno dalje gdje su četiri filtera osigurana.
8. Postupak prema bilo kojem od patentnih zahtjeva 1 do 7 a) koji se vrši na temperaturi okoline; i/ili b) koji se vrši bez vorteksiranja; i/ili c) gdje biološki materijal iz kojeg će nukleinske kiseline koje sadrže DNK biti izolirane sadrži stanice, opcionalno gdje su stanice u obliku peleta.
9. Kit za ekstrakciju nukleinskih kiselina koje sadrže DNK iz stanica, pri čemu komplet sadrži u kombinaciji: (A) (i) vodenu otopina koji sadrži, sastoji se u suštini od, ili se sastoji od vode, litij klorida u koncentraciji od 0.7 do 0.9 mola po litru, EDTA ili natrijeve soli istog u koncentraciji od 5 do 15mM, i 0.9 do 1.1% po masi SDS, (ii) vodene otopine soli gvanidinija, (poželjno oko 2M), (iii) etanol, (iv) prvu tekućinu za ispiranje koja sadrži litij klorid u koncentraciji od 0.3 do 0.6M, pri čemu navedena prva tekućina sadrži etanol u količini od 45% do 55% po zapremini, (v) drugu tekućini za ispiranje koja sadrži barem 80% po zapremini etanola, pri čemu je ravnoteža ako je prisutna voda, (vi) elucijski pufer, i (vii) spin kolonu koja ima filter za imobilizaciju DNK; ILI (B) (i) vodenu otopina koji sadrži, sastoji se u suštini od, ili se sastoji od litij klorida u koncentraciji od 0.2 do 0.3 mola po litru, EDTA ili natrijeve soli istog u koncentraciji od 1 do 5 milimola po litru, SDS u količini od 0.2-0.4% po masi, soli gvanidinija koja ima koncentraciju od 0.6 do 0.7M i etanola u količini od 35-45% po zapremini, (ii) prvu tekućinu za ispiranje koja sadrži litij klorid u koncentraciji od 0.3 do 0.6M, pri čemu navedena prva tekućina sadrži etanol u količini od 45% do 55% po zapremini, (iii) drugu tekućinu za ispiranje koja sadrži barem 80% po zapremini etanola, pri čemu je ravnoteža ako je prisutna voda, (iv) elucijski pufer, i (v) spin kolonu koja ima filter za imobilizaciju DNK.
10. Komplet prema patentnom zahtjevu 9 gdje filter sadrži barem jednu ploču boro-silikatnih vlakana.
11. Komplet prema patentnom zahtjevu 10 gdje filter sadrži četiri ploče boro-silikatnih vlakana.
HRP20201452TT 2015-03-17 2020-09-10 Izoliranje nukleinskih kiselina HRP20201452T1 (hr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
GB201504459A GB201504459D0 (en) 2015-03-17 2015-03-17 Isolation of DNA
PCT/GB2016/050738 WO2016147004A1 (en) 2015-03-17 2016-03-17 Isolation of nucleic acids
EP16719888.6A EP3277809B1 (en) 2015-03-17 2016-03-17 Isolation of nucleic acids

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HRP20201452T1 true HRP20201452T1 (hr) 2020-12-25

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US (1) US10934539B2 (hr)
EP (1) EP3277809B1 (hr)
JP (1) JP6713007B2 (hr)
CN (1) CN107743521B (hr)
AU (1) AU2016231944B2 (hr)
CY (1) CY1123363T1 (hr)
DK (1) DK3277809T3 (hr)
ES (1) ES2819903T3 (hr)
GB (1) GB201504459D0 (hr)
HR (1) HRP20201452T1 (hr)
HU (1) HUE051568T2 (hr)
LT (1) LT3277809T (hr)
PL (1) PL3277809T3 (hr)
PT (1) PT3277809T (hr)
RS (1) RS60850B1 (hr)
SI (1) SI3277809T1 (hr)
WO (1) WO2016147004A1 (hr)

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US11015186B2 (en) 2016-09-27 2021-05-25 Abbott Molecular Inc. Maximizing DNA yield of blood specimens collected in rapid clot tubes
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GB2581489B (en) 2019-02-15 2021-02-24 Revolugen Ltd Purification method
DE102020135124A1 (de) 2020-12-30 2022-06-30 Aj Innuscreen Gmbh Verfahren und System zur schnellen Isolierung von Nukleinsäuren direkt aus Vollblutproben

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GB201415674D0 (en) 2014-09-04 2014-10-22 Moorlodge Biotech Ventures Ltd Nucleic acid analysis

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EP3277809A1 (en) 2018-02-07
PT3277809T (pt) 2020-09-23
HUE051568T2 (hu) 2021-03-01
JP6713007B2 (ja) 2020-06-24
LT3277809T (lt) 2020-11-10
ES2819903T3 (es) 2021-04-19
RS60850B1 (sr) 2020-10-30
US20180066246A1 (en) 2018-03-08
AU2016231944B2 (en) 2021-07-29
AU2016231944A8 (en) 2017-12-07
US10934539B2 (en) 2021-03-02
PL3277809T3 (pl) 2020-11-16
SI3277809T1 (sl) 2020-11-30
CN107743521A (zh) 2018-02-27
GB201504459D0 (en) 2015-04-29
WO2016147004A1 (en) 2016-09-22
AU2016231944A1 (en) 2017-11-09
JP2018508239A (ja) 2018-03-29
DK3277809T3 (da) 2020-09-28
CN107743521B (zh) 2021-01-08
EP3277809B1 (en) 2020-06-24
CY1123363T1 (el) 2021-12-31

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