TIANcombi DNA Lyse & Det PCR Kit

Ho itlhoekisa ka potlako ha DNA ho tsoa lisebelisoa tse fapaneng bakeng sa ho sibolloa ha PCR.

TIANcombi DNA Lyse & Det PCR Kit e sebelisa moralo o ikhethileng oa ho paka o kenyelletsang li-reagents tsohle bakeng sa boitokiso bo potlakileng ba genomic DNA le kholiso ea PCR. E sebetsa bakeng sa tlhoekiso ea mohato o le mong oa DNA ea genome ho tsoa mehlala e fapaneng (lisele tsa semela, lipeo, lisele tsa liphoofolo, mali, tomoso le libaktheria) le kholiso le tlhahlobo ea PCR e latelang. Liprotheine, RNA le ho tlosoa ha metabolite ea bobeli, ho qhibiliha ha mokelikeli oa 'mele hammoho le mehato ea pula ha e hlokehe ts'ebetsong eohle ea tlhoekiso, ho etsa hore ts'ebetso e be bonolo ebile e potlake. Boleng ba sehlahisoa bo tsitsitse ebile boa tšepahala.

2 × Det PCR MasterMix e fanoeng ke sesebelisoa sena ke reagent e lumellanang haholo ea PCR e ka ntlafatsang ka nepo le ka kotloloho DNA ntle le tlhoko ea ho tlosa litšila tse kang liprotheine. Reagent ena e na le Taq DNA polymerase, dNTPs, MgCl₂, buffer, hammoho le ntlafatso, optimizer le stabilizer bakeng sa karabelo ea PCR. Ts'ebeliso ea reagent e etsa hore PCR e itšoare ka potlako, e bonolo, e bonolo, e ikhethang ebile e tsitsitse. Ka hona, kit ena e loketse haholo-holo ho hlahlojoa ka mokhoa o phahameng.

Katse. Che	Paka Size
4992527	20 ×l × 50 rxn
4992528	20 ×l × 200 rxn

Romela ya imeile ho re

Product Qaqileng

Mohlala oa Teko

LBH

Lihlahisoa tsa Product

Likaroloana

■ E bonolo ebile e potlakile: DNA e tsoang lithong tse fapaneng e ka ntšoa ka metsotso e 5 ntle le tlhoko ea ho sila haetrojene ea metsi.
■ Likopo tse pharalletseng: E sebelisoa bakeng sa makhasi a semela, peo, lisele tsa liphoofolo, lisampole tsa mali (mali a sa tsoa khuoa, anticoagulation, maqeba a mali, matheba a mali a omisitsoeng, jj.), Tomoso le libaktheria.
■ Ts'ebelisano e matla: Reagent ea PCR e loketse ntlafatso ea DNA e nkiloeng mehloling e fapaneng ea mehlala.

Likopo

■ Ho lemoha liphatsa tsa lefutso: Khetho e nepahetseng bakeng sa phumano e kholo ea liphatsa tsa lefutso.

Lintlha tsa bohlokoa

■ Bakeng sa disampole tse nang le maemo a phahameng a phenols, joalo ka makhasi a k'hothone, palo ea kenyelletso e lokela ho ba tlase ho 0,4 mg, ho seng joalo karabelo ea PCR e tla ameha.

Lihlahisoa tsohle li ka fetoloa bakeng sa ODM / OEM. Bakeng sa lintlha,ka kopo tlanya Customized Service (ODM / OEM)

E fetileng: K'hemik'hale ea lijalo tsa GMO le Kit ea Amplification

E 'ngoe: Kit ea PC Direct ea Mali

	DNA e ntšitsoe ho 5 mg ea makhasi le lipeo tsa poone, koro, raese, soya le k'hothone ka ho latellana. DNA e ntlafalitsoe ke PCR e sebelisa li-primer tse ikhethang. 6 μl DNA ho tsoa ho li-eluents tse 20 tsa μl li ile tsa laeloa ka tsela e ngoe le e ngoe. 1: taolo e nepahetseng genome; 2: tlohela mehlala; 3: mehlala ea peo; 4: NTC; 5: Litlhahlobo tsa D2000
	M: Letšoao la TIANGEN D2000; 1: Taolo e nepahetseng; 2-7: Palo ea libaka tse omisitsoeng tsa mali pampiring ea sefahla ke 1-6 ka tatellano; 8: Taolo e mpe. Phunya ea 3 mm e ne e sebelisetsoa ho nka matheba a mali a omisitsoeng ho tsoa pampiring ea sefahla e le sesebelisoa sa tlhahlobo ea tlhahiso. 6 μl DNA ho tsoa ho li-eluents tse 20 tsa μl li ile tsa laeloa ka tsela e ngoe le e ngoe.
	M: Letšoao la TIANGEN D2000; 1: Taolo e nepahetseng (genomic DNA e sebelisitsoe e le template); 2-7: Palo ea mali e ekelitsoeng ke 10 μl, 20 μl, 30 μl, 40 μl, 50 μl le 60 μl, ka ho latellana; 8-13: Palo ea mali e ekelitsoeng ke 10 μl, 20 μl, 30 μl, 40 μl, 50 μl le 60 μl, ka ho latellana; Mantaha 14: NTC. 6 μl DNA e tsoang ho li-eluents tse 20 tsa μl li ile tsa kenngoa ho gel ea agarose.

Q: Ha ho na lihlopha tsa matlafatso

Setšoantšo sa A-1

■ Template e na le litšila tsa protheine kapa li-inhibitors tsa Taq, jj. ——Tlhoekisa template ea DNA, tlosa litšila tsa protheine kapa u ntše template ea DNA ka lisebelisoa tsa tlhoekiso.

■ Tlatsetso ea template ha e felle ——Ho eketsa ka mokhoa o nepahetseng mocheso oa "denaturation" le ho lelefatsa nako ea denaturation.

■ Ho senyeha ha litempele ——Reka template hape.

Tlhahiso ea A-2

■ Boleng bo tlase ba li-primers ——Re kopanya hape selelekela.

■ Ho senyeha ha li-primer——Fana ka litlhahlobo tse phahameng tsa likhakanyo hore e be molumo o monyane bakeng sa ho li boloka. Qoba ho hatsela hangata le ho qhibiliha kapa nako e telele ea 4 ° C e bolokiloeng.

■ Moralo o sa nepahalang oa litlhahlobo (mohlala, bolelele ba lithuso ha boa lekana, botenya bo thehiloe lipakeng tsa lintho tsa pele, jj.

EA-3 Mg²⁺ho tsepamisa maikutlo

■ Mg²⁺ ho tsepamisa mohopolo ho tlase haholo ——Ho eketsa hantle Mg²⁺mahloriso: Lokisetsa Mg²⁺ mahloriso ka letoto la likarabelo ho tloha ho 1 mM ho isa ho 3 mM ka karohano ea 0.5 mM ho fumana Mg²⁺ mahloriso bakeng sa template le primer ka 'ngoe.

Mocheso oa ho koala oa A-4

■ Mocheso o phahameng oa ho koala o ama tlamo ea selelekela le template. Fokotsa mocheso o kenang 'me u ntlafatse boemo ka gradient ea 2 ° C.

Nako ea katoloso ea A-5

■ Nako e khuts'oane ea katoloso —- Eketsa nako ea katoloso.

P: Ho ba le tšoaetso ea bohata

Phenomena: Mehlala e mebe e bonts'a lihlopha tsa tatellano.

Tšilafalo ea A-1 ea PCR

■ Tšilafalo ea sefapano ea tatellano ea sepheo kapa lihlahisoa tsa ntlafatso ——Tlhokomelo hore o se ke oa tšela sampole e nang le tatellano ea sepheo sa sampuli e fosahetseng kapa oa e tšolla ka har'a tube ea centrifuge. Li-reagents kapa lisebelisoa li lokela ho etsoa mochini o ikemetseng ho felisa li-acid tsa nucleic, mme boteng ba tšilafalo bo lokela ho khethoa ka liteko tse mpe tsa taolo.

■ Tšilafalo ea li-reagent ——Qobella li-reagents ebe u li boloka ka mocheso o tlase.

A-2 Tona-khōlōr

■ Mg²⁺ ho tsepamisa mohopolo ho tlase haholo ——Ho eketsa hantle Mg²⁺ mahloriso: Lokisetsa Mg²⁺ mahloriso ka letoto la likarabelo ho tloha ho 1 mM ho isa ho 3 mM ka karohano ea 0.5 mM ho fumana Mg²⁺ mahloriso bakeng sa template le primer ka 'ngoe.

■ Moralo oa semmuso o sa nepahalang, 'me tatellano ea sepheo e na le thuto ea bongoli le tatellano e sa amaneng le sepheo. ——Litlhahlobo tsa pele-pele.

Q: Kholiso e sa tobang

Phenomena: Lihlopha tsa kholiso ea PCR ha li lumellane le boholo bo lebelletsoeng, ebang ke bo boholo kapa bo bonyenyane, kapa ka linako tse ling lihlopha tse ikhethileng tse matlafatsang le lihlopha tse sa ikhethileng tsa matlafatso lia etsahala.

Tlhahiso ea A-1

■ Lintlha tse qalang tsa semmuso

——Re-design ea khale.

■ Khakanyo ea semmuso e phahame haholo —— Ho eketsa mocheso oa denaturation hantle le ho lelefatsa nako ea denaturation.

Mofuta oa A-2 Mg²⁺ ho tsepamisa maikutlo

■ Mokha oa Mg²⁺ mahloriso a phahame haholo -— Fokotsa hantle mohopolo oa Mg2 +: Lokisetsa Mg²⁺ mahloriso ka letoto la likarabelo ho tloha ho 1 mM ho isa ho 3 mM ka karohano ea 0.5 mM ho fumana Mg²⁺ mahloriso bakeng sa template le primer ka 'ngoe.

A-3 Thermostable polymerase

■ Palo ea enzyme e fetelletseng —— Fokotsa palo ea enzyme ka nepo ka linako tse ling tsa 0.5 U.

Mocheso oa ho koala oa A-4

■ Mocheso o kenyang letsoho o tlase haholo —— Ho eketsa mocheso oa ho koala ka nepo kapa ho sebelisa mokhoa oa mekhahlelo e 'meli

Lipotoloho tsa A-5 PCR

■ Lipotoloho tse ngata tsa PCR ——Ho fokotsa palo ea lipotoloho tsa PCR.

P: Lihlopha tse hlabang kapa tse tlotsang

Tlhahiso ea A-1—— Boemo bo ikgethang ba maemo a mabe— - Iketsetse meralo ea mantlha, fetola boemo le bolelele ba semmuso ho ntlafatsa botumo ba eona; kapa ho etsa sehlaha sa PCR.

DNA ea A-2

——Thempleite ha e ea hloeka— - Hloekisa template kapa u ntše DNA e nang le lisebelisoa tsa tlhoekiso.

EA-3 Mg²⁺ ho tsepamisa maikutlo

——Mahl²⁺ ho tsepamisa mohopolo ho phahame haholo -— Fokotsa hantle Mg²⁺ mahloriso: Lokisetsa Mg²⁺ mahloriso ka letoto la likarabelo ho tloha ho 1 mM ho isa ho 3 mM ka karohano ea 0.5 mM ho fumana Mg²⁺ mahloriso bakeng sa template le primer ka 'ngoe.

E-4 dNTP

——Khokahano ea li-dNTP li holimo haholo —— Fokotsa mohopolo oa dNTP ka nepo

Mocheso oa Annealing oa 5

——Thempereichara e tlaase e kenang ——Ho eketsa hantle mocheso o tlatsetsang

Lipotoloho tsa A-6

—Kenale palo ea potoloho

P: Ke template ea bokae ea DNA e lokelang ho kenyelletsoa ho 50 μl PCR reaction system?

P: U ka holisa likaroloana tse telele joang?

Mohato oa pele ke ho khetha polymerase e loketseng. Kamehla Taq polymerase e ke ke ea bala liphoso ka lebaka la ho haelloa ke ts'ebetso ea 3'-5 'ea tokollo,' me ho se tšoane hantle ho tla fokotsa ts'ebetso ea likhechana. Ka hona, Taq polymerase e tloaelehileng e ke ke ea holisa hantle likaroloana tsa sepheo tse kholo ho feta 5 kb. Taq polymerase e nang le phetoho e khethehileng kapa botšepehi bo phahameng ba polymerase e lokela ho khethoa ho ntlafatsa katleho le ho fihlela litlhoko tsa sehlotšoana se selelele. Ntle le moo, kholiso ea likhechana tse telele e boetse e hloka phetoho e tsamaellanang ea moralo oa khale, nako ea denaturation, nako ea katoloso, pH ea buffer, jj. Hangata, li-primers tse nang le 18-24 bp li ka lebisa ho chai e ngata. Bakeng sa ho thibela tšenyo ea template, nako ea denaturation ho 94 ° C e lokela ho fokotsoa ho isa ho 30 sec kapa ka tlase ho potoloho, mme nako ea ho nyoloha mocheso ho ea ho 94 ° C pele amplification e lokela ho ba tlase ho 1 min. Ho feta moo, ho beha mocheso oa katoloso ka 68 ° C le ho rala nako ea katoloso ho latela sekhahla sa 1 kb / min ho ka netefatsa kholiso e sebetsang ea likhechana tse telele.

Q: U ka ntlafatsa ts'epahalo ea kholiso ea PCR joang?

Sekhahla sa phoso ea kholiso ea PCR se ka fokotsoa ka ho sebelisa li-polymerase tse fapaneng tsa DNA ka bots'epehi bo phahameng. Har'a li-polymerase tsohle tsa Taq DNA tse fumanehang ho fihlela joale, enzyme ea Pfu e na le sekhahla se tlase sa phoso le botšepehi bo phahameng ho fetisisa (bona tafole e hoketsoeng). Ntle le khetho ea enzyme, bafuputsi ba ka tsoela pele ho fokotsa sekhahla sa phetoho ea PCR ka ho ntlafatsa maemo a karabelo, ho kenyeletsoa le ho ntlafatsa sebopeho sa buffer, polasetiki ea thermostable polymerase le ho ntlafatsa palo ea potoloho ea PCR.

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