2 × HiF Taq wongeyeho Master mix
Injangwe No: HCR2014B
HIF Taq wongeyeho Master Mix (Hamwe Irangi) nigiteguye-gukoresha-2 solution igisubizo cyibanze kirimo Plus HIF ADN Polymerase, DNTPs, hamwe na buffer nziza.Antibodiyumu ebyiri za monoclonal mubushyuhe bwicyumba kibuza ibikorwa bya polymerase na 3 ′ → 5′ibikorwa byongeweho byongeweho kuri master mix kugirango byoroshye kandi byihariye Hot Start PCR.Ikintu cyo kwaguka cyongeweho kuri master mix kugirango itange enzyme ubushobozi burebure bwo kongera imbaraga, uburebure bwa amplification burashobora kugera kuri kb 13, enzyme ifite 5 ′ → 3 ′ ibikorwa bya polymerase ya ADN na 3 ′ → 5 ′ ibikorwa bya exonuclease, ubudahemuka bwayo bukubye inshuro 83 ubwa Taq ADN polymerase, bikubye inshuro 9 ibya ADN polymerase isanzwe.Birakwiriye kwongerwaho inyandikorugero zoroshye, ibicuruzwa byongera imbaraga ni impera.
2 × HIF Taq wongeyeho Master mix (Hamwe n'irangi) ifite ibyiza byo kwihuta kandi byoroshye, ibyiyumvo bihanitse, umwihariko ukomeye, ituze ryiza, nibindi, sisitemu yo kubyitwaramo ikeneye gusa kongeramo primers na templates, kandi irashobora kongerwaho na bibiri- intambwe protocole, koroshya intambwe zigeragezwa no kubika umwanya.Ibicuruzwa birimo amarangi yerekana amashanyarazi, kandi ibicuruzwa bya PCR birashobora gukoreshwa muburyo butaziguye kuri electrophorei.Mubyongeyeho, ibicuruzwa birimo kandi ibintu byihariye birinda, kugirango master mix irashobora gukomeza ibikorwa bihamye nyuma yo gukonjeshwa inshuro nyinshi.
Ububiko
Ibicuruzwa bigomba kubikwa kuri -25 ~ -15 ℃ kumwaka 1.
Ibisobanuro
| Ibisobanuro ku bicuruzwa | Master mix |
| Kwibanda | 2 × |
| Intangiriro ishyushye | Byubatswe muri Hot Itangira |
| Kurenga | Wibeshya |
| Umuvuduko wo kwitwara | Byihuta |
| Ingano (Igicuruzwa cyanyuma) | Kugera kuri 13kb |
| Ibisabwa mu bwikorezi | Urubura rwumye |
| Ubwoko bwibicuruzwa | Ubudahemuka bukabije PCR yibanze |
Amabwiriza
1.Sisitemu yo Kwitabira PCR
| Ibigize | Umubumbe (μL) |
| Icyitegererezo cya ADN | Birakwiriye |
| Imbere ya primer (10 μ mol / L) | 2.5 |
| Subiza Primer (10 μ mol / L) | 2.5 |
| 2 × HIF Taq wongeyeho Master mix | 25 |
| ddH2O | kugeza kuri 50 |
2.Basabwe gukoresha inyandikorugero zitandukanye
| Ubwoko bw'icyitegererezo | Ongera ibice kuva 1kb kugeza 10 kb |
| ADN rusange | 50ng-200 ng |
| ADN ya plasma cyangwa virusi | 10pg-20ng |
| cDNA | 1-2.5 µL (Ntukarenge 10% yumubare wanyuma wa PCR) |
3.Amasezerano yo kongera imbaraga
1) Intambwe ebyiri-Porotokole (inyandikorugero igoye)
| Intambwe | Ubushuhe. | Igihe | Amagare |
| Gutandukana kwambere | 98 ℃ | Imin | 1 |
| Gutandukana | 98 ℃ | 10sec | 30-35 |
| Kwagura | 68 ℃ | 30 amasegonda / kb | |
| Kwagura kwa nyuma | 72 ℃ | Imin | 1 |
2) Intambwe eshatu-Porotokole (protocole isanzwe)
| Intambwe | Ubushuhe. | Igihe | Amagare |
| Gutandukana kwambere | 98 ℃ | Imin | 1 |
| Gutandukana | 98 ℃ | 10sec | 30-35 |
| Annealing | 60 ℃ | 20 amasegonda | |
| Kwagura | 72 ℃ | 30 amasegonda / kb | |
| Kwagura kwa nyuma | 72 ℃ | Imin | 1 |
3) Annealing Gradient Protocol (inyandikorugero igoye)
| Intambwe | Ubushyuhe | Igihe | Amagare |
| Gutandukana kwambere | 98 ℃ | Imin | 1 |
| Gutandukana | 98 ℃ | Amasegonda 10 | 15 (1 ℃ kugabanuka kuri buri cyiciro) |
| Gradeent annealing | 70-55 ℃ | 20 amasegonda | |
| Kwagura | 72 ℃ | 30 amasegonda / kb | |
| Gutandukana | 98 ℃ | Amasegonda 10 |
20 |
| Annealing | 55 ℃ | 20 amasegonda | |
| Kwagura | 72 ℃ | 30 amasegonda / kb | |
| Kwagura kwa nyuma | 72 ℃ | Imin | 1 |
Ibiranga munsi ya amplification protocole itandukanye
| Porotokolel | Intambwe ebyiri | Intambwe eshatu | Gradeent annealing |
| Kugaragara. | byihuse | giciriritse | gahoro |
| Umwihariko | muremure | giciriritse | muremure |
| Umusaruro wa PCR | giciriritse | muremure | giciriritse |
| Igipimo cyo gutahura | muremure | giciriritse | muremure |
Inyandiko
Nyamuneka nyamuneka wambare PPE ikenewe, ikote rya laboratoire hamwe na gants, kugirango ubuzima bwawe n'umutekano wawe!
