Imwe Nhanho RT-qPCR SYBR Green Premix
Katsi Nhamba: HCB5140A
Imwe Nhanho RT-qPCR Syber Green Premix ndeye fluorescence quantification yakavakirwa paSYBR Green I dhayi.Kushandisa gene-specific primers, reverse transcription uye qPCR reactions zvinopedzwa muchubhu imwe, zvichibvisa kudiwa kwekudzokororwa kuvhura kepu-kuvhura uye mapipeting mashandiro, kunatsiridza zvakanyanya kushanda kwekuongorora uye kuderedza njodzi yekusvibiswa.Kumasamples eRNA, kit inoshandisa inodzivirira kupisa Reverse Transcriptase kune inoshanda cDNA synthesis uye HotStart Taq DNA Polymerase yekuwedzeredza kwehuwandu.Pasi peyakagadziridzwa buffer system, kunzwisiswa kwekiti kunogona kunge kwakakwira se0.1 pg kune yakanyatso kuratidzwa zvinangwa uye yakakwira se1 pg yezvinangwa zvakaratidzwa zvine mwero.Iyo kit inokodzera kukwidziridzwa uye quantification yeDNA samples.Iyo inogonesa kucherechedzwa kwekuonekwa uye quantification ye nucleic acids kubva kwakasiyana zvirimwa nemhuka samples, maseru uye tupukanana.
Zvikamu
| No | Zita | Volume | Volume |
| 1 | Advanced Buffer | 250 μL | 2×1.25 mL |
| 2 | Yepamberi Enzyme Mix | 20 μL | 200 μL |
| 3 | RNase Yemahara H2O | 250 μL | 2×1.25 mL |
Storage Conditions
Ichi chigadzirwa chinofanira kuchengetwa pa -25 ~ -15 ℃ kure nechiedza kwegore 1.
Mirayiridzo
1.Reaction system kugadzirisad
| Zvikamu | Vhoriyamu (μL) | Vhoriyamu (μL) | Final concentration |
| Advanced Buffer | 12.5 | 25 | 1× |
| Yepamberi Enzyme Mix | 1 | 2 | - |
| Pamberi Pekutanga (10 μmol/L)a | 0.5 | 1 | 0.2 μmol/L |
| Reverse Primer (10 μmol/L)a | 0.5 | 1 | 0.2 μmol/L |
| RNA Tamplateb | X | X | - |
| RNase Yemahara H2Oc | ku25 | kusvika ku50 | - |
Notes:
1) a.Tiye yekupedzisira primer concentration yaive 0.2 μmol/L, iyo inogonawo kugadziriswa pakati pe0.1 uye 1μmol/L sezvakakodzera.
2) b.Iyo reagent inonyanya kunzwisiswa, ine Total RNA muhuwandu hwe1pg-1μg, uye kuongororwa kwemasampuli evanhu kwakaratidza kupinza kwakakwana kwe1 pg-100 ng, kutonga kwehuwandu hweCt muhuwandu hwe15-30 sezvakakodzera.
3) c.Zvinokurudzirwa kushandisa 20μL kana 50μL kuve nechokwadi chechokwadi uye kuberekazve kwechinangwa chekusimudzira gene.
4) d.Ndapota gadzirirai mu-ultra-clean bhenji uye shandisai nuclease residue-free tips uye reaction tubes;mazano ane sefa cartridges anokurudzirwa.Dzivisa kusvibiswa kwemuchinjikwa uye kusvibiswa kweaerosol.
2.Reaction program
| Cycle step | Temp. | Nguva | Cycles |
| Reverse transcription | 50 ℃a | 6mins | 1 |
| Initial denaturation | 95℃ | 5 mins | 1 |
| Amplification reaction | 95℃ | 15 sec | 40 |
| 60 ℃b | 30 sec | ||
| Melting curve stage | Zviridzwa Defaults | 1 | |
Notes:
1) a.Iko reverse transcript tembiricha inogona kusarudzwa pakati pe50-55 ° C zvichienderana nekuyedza zvinodiwa.Kune sampuli dzeDNA, iyo reverse transcription process inogona kusiiwa.
2) b.Muzviitiko zvakakosha iyo annealing / yekuwedzera tembiricha inogona kugadziridzwa zvichienderana neyekutanga Tm kukosha, 60 ° C inokurudzirwa.
Notes
1. Ichi chigadzirwa ndechekutsvaga kushandiswa chete.
2. Ndokumbira ushande nemajasi erabhu uye magirovhosi anoraswa, kuitira kuchengeteka kwako.
