Buccal Swab/FTA Card DNA Isolation Kit Genomic DNA Extraction or Purifiction Kit from Buccal Swabs
Tlhaloso ea Kit:
Ka potlako hloekisa DNA ea boleng bo holimo ho tsoa lisampoleng tsa buccal swab/FTA Card.
◮Ha ho tšilafalo ea RNase:DNA-Feela Column e fanoeng ke kit e etsa hore ho khonehe ho tlosa RNA ho DNA ea genomic ntle le ho eketsa RNase nakong ea teko, ho thibela laboratori hore e se ke ea silafatsoa ke RNase e sa tloaelehang.
◮Lebelo le potlakileng:Foregene Protease e na le ts'ebetso e phahameng ho feta li-protease tse tšoanang, 'me e sila lisampole tsa linama kapele;ts'ebetso e bonolo, 'me ts'ebetso ea ho ntša DNA ea genomic e ka phethoa nakong ea metsotso ea 20-80.
◮E loketse:The centrifugation e etsoa ka mocheso oa kamore, 'me ha ho hlokahale 4 ° C mocheso o tlase oa centrifugation kapa ethanol precipitation ea DNA.
◮Tšireletseho:Ha ho hlokahale ho ntšoa ha reagent ea organic.
◮Boleng bo phahameng:DNA e nkiloeng e na le likhechana tse kholo, ha e na RNA, ha e na RNase, le li-ion tse tlase haholo, tse ka fihlelang litlhoko tsa liteko tse fapaneng.
◮Sistimi ea Micro-elution:E ka eketsa bongata ba DNA ea genomic, e leng bonolo ho lemoha kapa ho etsa liteko.
Tlhaloso
Setsi sena se fana ka mokhoa o sebetsang hantle le o potlakileng oa ho fumana DNA ea genomic e phahameng ea concentration ho tloha buccal swabs le FTA Card (matheba a mali).Ho sebelisa k'hamphani ea rona's e ikhethang ea DNA-feela silica membrane spin column le formula, e kopantsoeng le Foregene Protease, DNA e phahameng ea concentration, ea boleng bo phahameng ea genomic e ka ntšoa ka metsotso e 80.Kholomo e nyane e etselitsoeng ho hloekisa e kopanya DNA ea genomic, 'me DNA e ka hlakoloa ka chelete e nyane.15μl) ho eketsa bongata ba DNA e fumanoeng ea genomic, e loketseng ho fumanoa kapa ho etsa liteko.Kit e ka sebetsana le sampuli e le 'ngoe kapa ho feta ka nako,' me ts'ebetso ea tlhoekiso ha e hloke ho ntšoa ha lintho tse phelang tse kang phenol, chloroform, le nako e jang isopropanol kapa ethanol precipitation, 'me ts'ebetso e bonolo ebile e boloka nako.
Litlhaloso
50 Preps
Likarolo tsa Kit
| Sekhahla sa ST1 |
| Sekhahla sa ST2 |
| Linear Acrylamide |
| Sekhahla sa PW |
| Buffer WB |
| Buffer EB |
| Foregene Protease |
| DNA-Feela Kholomo |
| Litaelo |
Likarolo le melemo
-Ha ho tšilafalo ea RNase: Kholomo ea DNA-Feela e fanoeng ke kit e etsa hore ho khonehe ho tlosa RNA ho DNA ea genomic ntle le ho eketsa RNase nakong ea teko, ho qoba laboratori hore e se ke ea silafatsoa ke RNase e sa tloaelehang.
-Lebelo le potlakileng: Foregene Protease e na le ts'ebetso e phahameng ho feta li-protease tse tšoanang, e sila lisampole kapele;tshebetso e bonolo.
-E bonolo: The centrifugation e etsoa ka mocheso oa kamore, 'me ha ho hlokahale 4 ° C mocheso o tlaase oa centrifugation kapa ethanol precipitation ea DNA.
-Tshireletso: Ha ho hlokahale ho ntšoa ha reagent ea organic.
-Boleng bo phahameng: DNA e nkiloeng ea genomic e na le likhechana tse kholo, ha e na RNA, ha e na RNase, le li-ion tse tlaase haholo, tse ka finyellang litlhoko tsa liteko tse sa tšoaneng.
-Micro-elution system: E ka eketsa bongata ba DNA ea genomic, e leng bonolo bakeng sa ho lemoha kapa ho etsa liteko.
Sesebelisoa sa Kit
E loketse ho hloekisa DNA ea genomic ho tloha ho mehlala e latelang: li-buccal swabs, FTA Card (mabala a mali).
Storage le Shelf bophelo
-Setsi sena se ka bolokoa likhoeli tse 12 tlas'a maemo a omileng mocheso oa kamore (15-25 ° C);haeba e hloka ho bolokoa nako e telele, e ka bolokoa ho 2-8°C.
Tlhokomeliso: Haeba e bolokiloe mochesong o tlase, tharollo e atisa ho ba le pula.Pele o sebelisoa, etsa bonnete ba hore o beha tharollo ka har'a kit mocheso oa kamore ka nako e itseng.Haeba ho hlokahala, e chese pele ka ho hlapa metsi a 37 ° C ka metsotso e 10 ho qhaqha precipitate, 'me e kopanye pele e sebelisoa.
-Forene Protease tharollo e na le mokhoa o ikhethang, o sebetsang ha o bolokiloe mocheso oa kamore nako e telele (likhoeli tse 3);mosebetsi oa eona le botsitso li tla ba molemo ha li bolokiloe ho 4 ° C, kahoo ho kgothaletswa ho e boloka ho 4 ° C, hopola hore u se ke ua e boloka ho -20 ° C.
Kholomo ea tlhoekiso e koalehile
Ka kit ena, ts'ebetsong ea ts'ebetso ea DNA ea genomic, kholomo ea tlhoekiso e adsorbed ka ho toba ka sampuli ea enzymatic lysis motsoako ntle le mohato oa centrifugation, 'me mohala oa tlhoekiso o ka thibeloa ka lebaka la enzymization e sa fellang le viscosity e phahameng ea sampuli.
Lisosa tse latelang tse ka hlahang ke tse latelang:
1. Tshilo e sa fellang ya enzymatic ya disampole tsa dithishu.
Keletso: Nako ea ts'ebetso ea sampole ea Foregene Protease e ka atolosoa ka nepo kapa supernatant e ka nkuoa kamora centrifugation ka 12,000 rpm (~13,400 × g) bakeng sa 5 min.
2. Tšebeliso e feteletseng ea lisampole tsa linama kapa lisele tse kholo.
Keletso: Ho molemo hore u se ke ua feta 1 Buccal swab ka sampuli;haeba sampole e le kholo haholo, eketsa litekanyetso tsa Buffer ST1, Foregene Protease, buffer ST2 ka nepo.
3. Mohlala oa viscosity o phahame haholo.
Keletso: lisampole li ka hlapolloa ka mokhoa o nepahetseng ka 10 mM ea Tris-HCl pele ho ho ntšoa ha genomic DNA.
4. Likhechana tsa karete ea mali li anngoe.
Keletso: Nako ea nakoana ea centrifugation ea mohato oa 6 oa sebaka sa mali (FTA Card) genomic extraction e ka atolosoa ka nepo.
Kotulo e tlase kapa ha e na DNA
Hangata ho na le lintlha tse fapaneng tse amang tlhahiso ea DNA ea genomic, ho kenyelletsa le tšimoloho ea mohlala, maemo a polokelo ea sampole, ho lokisoa ha mohlala, ho qhekella, joalo-joalo.
Genomic DNA e ke ke ea fumanoa nakong ea ho ntšoa
Lisosa tse ka bang teng ke tse latelang:
1. Paballo e fosahetseng ea lisampole kapa polokelo nako e telele haholo e lebisa ho senyeha ha DNA ea genomic.
Keletso: Li-swabs tsa molomo ka ho khetheha li lokela ho etsoa sampole e ncha, 'me ha ho bohlale ho sebelisa li-swabs tse bolokiloeng bakeng sa ts'ebetso ea ho ntša DNA ea genomic;disampole tsa letheba la madi di lokela ho etsa bonnete ba hore boleng bo tshwaneleha mme nako ya polokelo ha ya lokela ho ba telele haholo.
2. Tšebeliso e fokolang haholo ea lisele e ka 'na ea etsa hore ho se ke ha ntšoa DNA e tsamaisanang le eona.
Keletso: Latela litaelo tsa lisampole tsa buccal swab tataisong ea ts'ebetso, 'me u hlakole ka makhetlo a mangata kamoo ho ka khonehang e le hore lisele tse lekaneng li ka hokeloa ho oral swab bakeng sa ho ntšoa ha DNA ea genomic;bakeng sa ho ntšoa ha letheba la mali, sebaka sa ho seha letheba la mali se ka eketsoa ka nepo.
3. Foregene Protease e bolokiloe ka mokhoa o sa lokelang, e leng se bakang ho fokotseha ha mosebetsi oa eona kapa ho se sebetse.
Keletso: Netefatsa maemo a polokelo ea Foregene Protease kapa sebakeng sa eona ka Foregene Protease e ncha bakeng sa karabelo ea enzymatic.
4. Ho boloka nako e sa lokelang ea kit kapa nako ea ho boloka nako e telele haholo, ho fella ka ho hlōleha ha likarolo tse ling ka har'a kit.
Keletso: Reka sesebelisoa se secha sa ho itšehla thajana sa Buccal swab DNA bakeng sa lits'ebetso tse amanang.
5. Buffer WB ha e kenye ethanol e felletseng.
Keletso: Netefatsa hore buffer WB e eketsa molumo o nepahetseng oa ethanol e felletseng.
6. Sehlahisoa ha sea kenyelletsoa ka nepo filiming ea silicone.
Keletso: Eketsa marotholi a 65 ° C a futhumetseng pele a futhumala bohareng ba lera la silicone 'me u siee mocheso oa kamore metsotso e 5 ho eketsa ts'ebetso ea tlhahiso.
DNA ea lihlahisoa tse tlaase tsa genomic e arotsoe
Lisosa tse latelang tse ka hlahang ke tse latelang:
1. Paballo e fosahetseng ea lisampole kapa polokelo nako e telele haholo e lebisa ho senyeha ha DNA ea genomic.
Keletso: Li-swabs tsa molomo li ka khethoa e le tse sa tsoa etsoa sampole, 'me li-swabs tse bolokiloeng ha lia lokela ho sebelisoa bakeng sa ho ntšoa ha DNA ea genomic.
2. Haeba palo ea sampole ea linama e nyane haholo, litaba tsa DNA tse nkiloeng li tla fokotseha.
Keletso: Latela litaelo tsa oral swab tsa sampole bukeng ea ts'ebetso, u hlakole ka makhetlo a mangata kamoo ho ka khonehang e le hore lisele tse lekaneng li ka hokeloa ho oral swab bakeng sa ho ntšoa ha genomic DNA.
3. Foregene Protease e bolokiloe ka mokhoa o sa lokelang, e leng se bakang ho fokotseha ha mosebetsi oa eona kapa ho se sebetse.
Keletso: Netefatsa maemo a polokelo ea Foregene Protease kapa sebakeng sa eona ka Foregene Protease e ncha bakeng sa karabelo ea enzymatic.
4. Eluent mathata.
Keletso: Sebelisa Buffer EB bakeng sa elution;haeba o sebelisa ddH2O kapa lintlha tse ling, tiisa hore pH ea eluate e pakeng tsa 7.0-8.5.
5. The eluate ha e eketsoe ka nepo dropwise.
Keletso: Eketsa marotholi a seng makae bohareng ba lera la silicone 'me u siee mocheso oa kamore metsotso e 5 ho eketsa katleho ea elution.
6. Mokelikeli oa elution o bokellana hanyenyane haholo.
Keletso: Sebelisa eluent bakeng sa genomic DNA elution joalo ka ha ho hlokoa litaelong, bonyane e seng ka tlase ho 15 μl.
Bohloeki bo tlase ba DNA ea genomic bo arohane
Bohloeki bo tlase ba DNA ea genomic bo ka lebisa ho hloleheng kapa liphello tse sa khotsofatseng tsa liteko tse tlaase, tse kang: li-enzyme li ke ke tsa khaoloa li bulehile, PCR ha e khone ho fumana sekhechana sa liphatsa tsa lefutso, joalo-joalo.
Lisosa tse ka bang teng ke tse latelang:
1. Tšilafalo ea Heteroprotein, tšilafalo ea RNA.
Tlhahlobo: Kholomo ea tlhoekiso ha ea ka ea hlatsuoa ho sebelisoa Buffer PW;kholomo ea tlhoekiso ea Buffer PW ha ea ka ea hlatsuoa ho sebelisoa lebelo le nepahetseng la centrifugal.
Keletso: Etsa bonnete ba hore ha ho na pula ho matla a phahametseng tlhaho pele u eketsa ethanol;etsa bonnete ba ho hlatsoa kholomo ea tlhoekiso ho latela litaelo, 'me mohato ona o ke ke oa tloheloa.
2. Tšilafalo ea ion tšilafalo.
Tlhahlobo: Kholomo ea tlhoekiso ea li-buffer WB e siiloe kapa ea hlatsuoa hanngoe feela, e leng se ileng sa baka tšilafalo e setseng ea ionic.
Keletso: Etsa bonnete ba hore u hlatsoa Buffer WB makhetlo a 2 kamoo u laetsoeng ho tlosa li-ion tse setseng ka hohle kamoo ho ka khonehang.
3. Tšilafalo ea enzyme ea RNA.
Analysis: Li-RNA tsa kantle ho naha li kentsoe ho buffer;Ts'ebetso ea ho hlatsoa PW ea Buffer e ne e fosahetse, e bakileng masala a RNase, a ama ts'ebetso ea liteko tsa RNA e tlase, joalo ka mongolo oa in vitro.
Keletso: Foregene series nucleic acid isolation kits e ka tlosa RNA ntle le tlatsetso ea RNase, ka hona buccal Swab/FTA Card DNA Isolation Kit ha e hloke ho eketsa RNase;etsa bonnete ba hore u latela litaelo tsa kholomo ea tlhoekiso ea ho hlatsuoa ea Buffer PW, 'me mohato ona o ke ke oa tloheloa.
4. Masala a Ethanol.
Tlhahlobo: Buffer WB ha ea ka ea etsa centrifugation e se nang letho ka mor'a ho hlatsoa kholomo ea tlhoekiso.
Keletso: Etsa ts'ebetso e nepahetseng ea tube centrifugation e nepahetseng ho latela litaelo.
5. Tšilafalo e ’ngoe ea litšila.
Tlhahlobo: Lisampole tse bolokiloeng kapa lisampole tse khethehileng ha li etsoe esale pele.
Keletso: Hlophisa sampole esale pele joalokaha u laetsoe.
