Ngqo RT-qPCR Kit II
Inkcazelo yeKit:
◮Ngqo RT-qPCR Kit, usebenzisa Foreasy Reverse Transcriptase kunye Foreasy HS Taq DNA Polymerase ephuhliswayo yi Foregene, kunye buffer reaction unique, yenza le khithi ukuxhathisa olomeleleyo kunye nokuhambelana, kwaye inokuvavanya ngokuthe ngqo reaction ukusebenzisa inkqubo Foregene Lysis njenge template.Le khithi inokufaka isicelo sekhithi yokufumanisa i-COVID-19 nucleic acid kunye nezinye iibhaktheriya ze-pathogenic ze-nucleic acid yophuhliso lwekhithi yokufumanisa.
Inkcazo
Direct RT-qPCR Kit ibonelela ngokwahlukeneyo RT-qPCR buffer, esebenza kakuhle Foreasy HS Taq DNA Polymerasekunye ne-Foreasy Reverse Transcriptase(MMLV),ukwenza kube lula ekudalweni kweekhithi zomva kunye nohlengahlengiso lwenkqubo, ngaphakathi kokuqinisekiswa okulula.
Ngqo RT-qPCR Kit, usebenzisa Foreasy Reverse Transcriptase kunye Foreasy HS Taq DNA Polymerase ephuhliswayo yi Foregene, kunye buffer reaction unique, yenza le khithi ukuxhathisa olomeleleyo kunye nokuhambelana, kwaye inokuvavanya ngokuthe ngqo reaction ukusebenzisa inkqubo Foregene Lysis njenge template.Le khithi inokufaka isicelo sekhithi yokufumanisa i-COVID-19 nucleic acid kunye nezinye iibhaktheriya ze-pathogenic ze-nucleic acid yophuhliso lwekhithi yokufumanisa.
Le kit ingaba yinxalenye yemveliso ye-IVD, isebenzisa ngokulula nangokukhawuleza.Ifuna kuphela ukuqinisekiswa okulula, ngaphandle kokuphuhlisa kwakhona.
Iinkcukacha
500T, 50,000T
Amacandelo ekhithi
| Imixholo yekhithi (20 μL inkqubo yokusabela) | IM-05111-02 | IM-05112-02 |
| 500 T | 50,000 T | |
| I-Foreasy Reverse Transcriptase (200 U/μL) | 50 μL×1 | 1 ml×5 |
| Foreasy HS Taq DNA Polymerase (5 U/μL) | 100 μL×1 | 1 ml×10 |
| I-2 × I-RT-qPCR ngqoIsithinteli | 1 mL ×5 | 500 mL ×1 |
| Umyalelo | 1 | 1 |
Amanyathelo okusebenza
A: Lungiselela itemplate kunye ne-reagent
1. Lungiselela i-template ye-RNA elungiselelwe (Kucetyiswa ukuba usebenzise i-Foregene Total RNA Isolation Kit series kit ukuze ukhuphe kwaye uhlambulule i-template ye-RNA) okanye isampula yokuqhawula imveliso (Kucetyiswa ukuba kusetyenziswe inkqubo ye-Foregene Lysis), ii-primers ezithile (10 μM) kunye nezinye izinto ezisetyenziswayo ezifanelekileyo, isixhobo.
2. Beka i-2 × Direct RT-qPCR Buffer, RNase-Free ddH2O kunye ne-20× ROX Reference Dye (ukuba iyafuneka) kwibhokisi enomkhenkce, okwenza ezi zinyibilike ngokwemvelo, kwaye zixutywe ngobunono.
B: Lungisa inkqubo ye-RT-qPCR
Fumana isiqingatha somthamo we-reaction buffer yenkqubo yokusabela (umzekelo, ukuba umthamo wenkqubo ngama-20 μL, kufuneka ufumane i-10 μL 2× Direct RT-qPCR Buffer). Kubainani elihambelanayo le-enzyme, sicebisa ukubaM-MLV: 10-30 U/20 μL inkqubo yokusabela,TaqDNA polymerase:1-2 U / 20 μL inkqubo yokusabelale yingcebiso yethu, kufuneka uyivavanye kwaye uyilungelelanise), yongeza ithemplate ye-RNA, ii-primers ezithile kunye ne-probes, kwaye ungeze i-ddH ye-RNase-Free2O ukuya kuma-20 μL.Ulungiselelo oluthile lwenkqubo yokusabela ye-RT-qPCR ingabhekiswa kule tafile ilandelayo 1.
Itheyibhile 1 : Ukulungiswa kwenkqubo ye-RT-qPCR
| Icandelo | Umthamo | Ugxininiso lokugqibela |
| 2× Ngqo RT-qPCR Buffer | 10 μL | 1× |
| I-Foreasy Reverse Transcriptase(MMLV) | 10-30 U | |
| Foreasy HS Taq DNA Polymerase | 1-2 U | |
| I-Primer Phambili(10 μM) | 0.8 μL | 50-900nM |
| Reverse Primer(10 μM) | 0.8 μL | 50-900nM |
| Probe(4 μM) | 1 μL | 200nM |
| Template(RNA okanye Lysate) | X μL | |
| I-20 × i-ROX ye-Reference Dye | - | 1* |
| RNase-FreeddH2O | (6.4-X) μL | |
| Umthamo opheleleyo | 20 μL |
Qaphela: I-Primer yokuPhambili kunye ne-Reverse Primer ziziiprayimari ezithile zejini ekujoliswe kuyo.Inkqubo ye-qPCR inokulungelelaniswa ngokovavanyo olusebenzayo kunye neemodeli ze-PCR.Sincoma i-400nM yogxininiso lokugqibela lweziqalo ezininzi.Lungisa umthamo wee-primers ezithile kunye ne-probes ngokogxininiso olulungisiweyo kunye nogxininiso lokugqibela olucetyiswayo, nceda.
I-1 *: Khetha ugxininiso olufanelekileyo lokugqibela lwe-ROX Reference Dye ngokwezixhobo ezahlukeneyo zePCR.Olona xinaniso lwe-ROX Reference Dye lwezixhobo eziqhelekileyo ze-PCR luboniswa kwitheyibhile ilandelayo:
| Izixhobo zePCR ezinobungakanani | Ugxininiso lokugqibela lwe-ROX Reference Dye |
| ABI PRISM 7000/7300/7700/7900HT/Inyathelo lokuqala,njl. | 1× (umzekelo, yongeza i-1μl 20×ROX Reference Dayi kwi-20 μL yokusabela inkqubo) |
| ABI 7500, 7500 Fast, Stratagene Mx3000P, Mx3005P kunye neMx4000, njl. | 0.5× (umzekelo, yongeza 0.5μl 20×ROX Reference Idayi kwi-20 μL yokusabela inkqubo) |
| Isixhobo se-PCR se-Roche, isixhobo se-Bio-Rad PCR, isixhobo se-PCR sobungakanani be-Eppendorf, njl. | Ngaphandle kwe-ROX Reference Daye |
Ugcino kunye nobomi beShelf
Ikhithi kufuneka igcinwe ku -20 ± 5℃.Kufuneka igcinwe ngoko nangoko kwi -20 ℃ efrijini ye-thermostatic.Ixesha lokuqinisekisa yiminyaka emi-2 ukuba igcinwe kwimeko efanelekileyo.
