Imwe Nhanho Yekukurumidza RT-qPCR Probe Premix-UNG
Katsi Nhamba: HCR5143A
Imwe Nhanho RT-qPCR Probe Kit (ZVEKUFANIRA) ndeye probe-based RT-qPCR inokurumidza kuona kit inokodzera single-plex kana multiplex quantitative PCR uchishandisa RNA setemplate (senge RNA hutachiona).Ichi chigadzirwa chinoshandisa chizvarwa chitsva cheantibody-yakagadziridzwa Taq DNA Polymerase uye imwe-nhanho yakatsaurirwa Reverse Transcriptase, ine yakagadziridzwa buffer yekukurumidza kukwidziridzwa, iyo inokurumidza kukwidziridza kumhanya, yakakwirira amplification kunyatsoita uye hunhu.Inotsigira kukwidziridzwa kwakaenzana mune ese-single-plex uye multiplex yeakaderera uye yakakwira yekumisikidza samples munguva pfupi.
Zvikamu
1. 5×RT-qPCR buffer (U+)
2. Musanganiswa we enzyme (U+)
Notes:
a.5 × RT-qPCR buffer (U+) inosanganisira dNTP uye Mg2+;
b.Enzyme mix (U+) inosanganisira reverse transcriptase, Hot Start Taq DNA polymerase, RNase inhibitor uye UDG;
c.Shandisa RNase-Yemahara matipi, EP machubhu, nezvimwe.
Usati washandisa, nyatso sanganisa iyo 5 × RT-qPCR buffer (U+).Kana paine chero kunaya mushure mekunyunguduka, mirira kuti buffer idzokere kune tembiricha yekamuri, sanganisa nekunyungudika, wobva wazvishandisa zvakajairika.
Storage Conditions
Chigadzirwa chacho chinotumirwa nechando chakaoma uye chinogona kuchengetwa pa -25 ~ -15 ℃ kwegore 1.
Mirayiridzo
1. Kuita System
| Zvikamu | Kuwanda (20 μL kuita) |
| 2 ×RT-qPCR buffer | 4μL |
| Musanganiswa we enzyme (U+) | 0.8μL |
| Primer Forward | 0.1~ 1.0μM |
| Primer Reverse | 0.1~ 1.0μM |
| TaqMan Probe | 0.05~0.25μM |
| Template | X μL |
| RNase-Isina Mvura | kusvika ku25μL |
Notes: React volume is 10-50μL.
2. Protocol yeCycling (Szvakajairika)
| Cycle step | Temp. | Nguva | Cycles |
| Reverse Transcription | 55 ℃ | 10 min | 1 |
| Kutanga Denaturation | 95 ℃ | 30 sec | 1 |
| Denaturation | 95 ℃ | 10 sec | 45 |
| Kuwedzera/Kuwedzera | 60 ℃ | 30 sec |
| Cycling Protocol (Kurumidza) Cycle step |
Temp. |
Nguva |
Cycles |
| Reverse Transcription | 55 ℃ | 5 min | 1 |
| Kutanga Denaturation | 95 ℃ | 5 p | 1 |
| Denaturation | 95 ℃ | 3 p | 43 |
| Kuwedzera/Kuwedzera | 60 ℃ | 10 p |
Notes:
a.Reverse transcript tembiricha iri pakati pe50 ℃ kusvika 60 ℃, kuwedzera tembiricha kunobatsira kukudza zvimiro zvakaoma uye yakakwirira CG yemukati matemplate;
b.Iyo yakakwana yekudziya tembiricha inoda kugadziridzwa zvichienderana neiyo Tm kukosha kweiyo primer, uye sarudza ipfupi nguva yekuunganidza fluorescence chiratidzo chinoenderana neChaiyo Nguva PCR chiridzwa.
Notes
Ndokumbira upfeke PPE inodiwa, jasi rakadaro uye magurovhosi, kuve nechokwadi chehutano hwako uye kuchengetedzeka!
