Multiplex Inyathelo elinye RT-qPCR Premix
Inkcazo
Inombolo yekati: HCR5141A
I-Multiplex One Step RT-qPCR Premix yi-multiplex quantitative PCR Kit esekelwe kwi-RNA njenge template.Kuvavanyo, ukukhutshelwa umva kunye ne-PCR yobungakanani zenziwa kwityhubhu yokusabela efanayo, ukwenza lula umsebenzi wokulinga kunye nokunciphisa umngcipheko wosulelo.Uyilo olulodwa lwe-buffer kunye nomxube we-enzyme lunokusetyenziswa kwinkqubo ye-lyophilized enenyathelo elinye.Ikhithi isebenzisa i-Reverse Transcriptase enganyangekiyo kubushushu ukuze idityaniswe ngokufanelekileyo ne-cDNA yomtya wokuqala kusetyenziswa i-hotstart Taq DNA Polymerase yokwandisa ubungakanani.Iqulethe isithinteli sokusabela esilungiselelweyo, umxube wee-enzymes njl.njl., kunye nezinto ezithintela ngokufanelekileyo ulwandiso lwe-PCR olungangqalanga kunye nokuphucula ukusebenza kakuhle kokwandiswa kweempendulo ze-qPCR ezininzi zongezwa, okwenza ukuba i-fluorescence yokwandisa ubungakanani be-multiple ngelixa iqinisekisa impumelelo yokukhulisa iiprimer.
Amacandelo
| Igama |
| 1. I-Lyo-Buffer |
| 2. Umxube weLyo-Enzyme |
| 3. Umkhuseli weLyo |
Umqathango wezothuthoion
A: I-Lyo-buffer kunye nomkhuseli: -25~-15℃, ubomi beshelufu ngunyaka omnye.
B: Umxube we-Lyo-enzyme, 2-8 ℃, ubomi beshelufu ziinyanga ezi-6.
Umyalelo wokusebenza
1. Inkqubo yokusabela (Thatha i-25μL njengomzekelo)
| Amacandelo | Umthamo (μL) | Ukugxininiswa kokugqibela |
| I-Lyo-Buffer | 6 | 1* |
| Umxube weLyo-Enzyme | 1 | - |
| Lyo-Mkhuseli | 8 | - |
| I-Primer Mix (10μM) | 1 | 0.1- 1uM |
| Probe Mix (10μM) | 0.5 | 0.05-0.5uM |
| Isakhelo seRNA | 5 | - |
| I-DEPC H2O | Ukuya kuthi ga kwi-25 | - |
2. IProtokholi yokuQhuba ibhayisekile
1) IProtokholi yoMgangatho weBhayisikile
| Inqanaba lokusabela | Ubushushu | Ixesha | Umjikelo | |
| 1 | Reverse transcription | 50°Ca | 10min | 1 |
| 2 | I-denaturation yokuqala | 95°C | 5min | 1 |
| 3 | Ukusabela kokukhulisa | 95°C | 15sec | 45 imijikelo |
| 60°Cb | 30secc |
2) IProtocol yokuHamba ngeBhayisikile ngokukhawuleza
|
| Inqanaba lokusabela | Ubushushu | Ixesha | Umjikelo |
| 1 | Reverse transcription | 50°Ca | 2 min | 1 |
| 2 | I-denaturation yokuqala | 95°C | 2 imizuzwana | 1 |
| 3 | Ukusabela kokukhulisa | 95°C | 1 umzuzw |
45 imijikelo |
| 60°Cb | 13secc |
Phawula:
a) Umva ushicilelo: Iqondo lokushisa linokukhetha i-42 ° C okanye i-50 ° C ngemizuzu eyi-10-15.
b) Ukusabela kokukhulisa: Ubushushu buhlengahlengiswa ngokwexabiso le-Tm lee-primers ezilungiselelwe.
c)I-Fluorescence uphawu ukufumana: Nceda usete inkqubo yovavanyo ngokweemfuno zeisixhobo manual.
Iinkcukacha zoBugcisa/iiNgcaciso
| Isiqalo esishushu | Isiqalo esishushu esakhelwe ngaphakathi |
| Indlela yokubona | Ukufunyanwa kwe-Primer-probe |
| Indlela yePCR | Inyathelo elinye RT-qPCR |
| Uhlobo lwesampulu | I-RNA |
Amanqaku
1. Le mveliso isetyenziselwa uphando kuphela.
2. Nceda unxibe iPPE eyimfuneko, idyasi yelebhu enjalo kunye neeglavu, ukuqinisekisa impilo yakho kunye nokhuseleko!
