Direct RT-qPCR Kit II
Kufotokozera Kwa Zida:
◮Direct RT-qPCR Kit, pogwiritsa ntchito Foreasy Reverse Transcriptase ndi Foreasy HS Taq DNA Polymerase yopangidwa ndi Foregene, yokhala ndi buffer yapadera, imapangitsa chida ichi kukhala cholimba komanso chogwirizana, ndipo chimatha kuyesa mwachindunji kugwiritsa ntchito kachitidwe ka Foregene Lysis ngati template.Zidazi zitha kuyika zida zozindikirira za COVID-19 nucleic acid ndi mabakiteriya ena a pathogenic nucleic acid kuzindikira zida.
Kufotokozera
Direct RT-qPCR Kit imapereka bafa ya RT-qPCR, yothandiza kwambiri ya Foreasy HS Taq DNA Polymerasendi Foreasy Reverse Transcriptase(MMLV),kupanga kukhala kosavuta kupanga zida zam'mbuyo ndi kusintha kwadongosolo, mkati mwa kutsimikizira kosavuta.
Direct RT-qPCR Kit, pogwiritsa ntchito Foreasy Reverse Transcriptase ndi Foreasy HS Taq DNA Polymerase yopangidwa ndi Foregene, yokhala ndi buffer yapadera, imapangitsa chida ichi kukhala cholimba komanso chogwirizana, ndipo chimatha kuyesa mwachindunji kugwiritsa ntchito kachitidwe ka Foregene Lysis ngati template.Zidazi zitha kuyika zida zozindikirira za COVID-19 nucleic acid ndi mabakiteriya ena a pathogenic nucleic acid kuzindikira zida.
Zida izi zitha kukhala gawo lazinthu za IVD, kugwiritsa ntchito mosavuta komanso mwachangu.Zimangofunika kutsimikizira kosavuta, osapanganso.
Zofotokozera
500T, 50,000T
Zida zamagulu
| Zamkatimu (20 μL reaction system) | IM-05111-02 | IM-05112-02 |
| 500 T | 50,000 T | |
| Foreasy RTranscriptase yonse (200 U /μL) | 50 μl pa×1 pa | 1 ml×5 pa |
| Foreasy HS Taq DNA Polymerase (5 U/μL) | 100 μL×1 pa | 1 ml× 10 pa |
| 2 × Direct RT-qPCRBafa | 1 mL × 5 | 500mL × 1 |
| Malangizo | 1 | 1 |
Njira Zogwirira Ntchito
A: Konzani template ndi reagent
1. Konzani template ya RNA yokonzekera (Ndibwino kuti mugwiritse ntchito zida za Foregene Total RNA Isolation Kit kuti muchotse ndi kuyeretsa template ya RNA) kapena chitsanzo chophwanyidwa (Ndikulimbikitsidwa kugwiritsa ntchito dongosolo la Foregene Lysis), zoyambira zenizeni (10 μM) ndi zina zowonjezera, chida.
2. Ikani 2 × Direct RT-qPCR Buffer, RNase-Free ddH2O ndi 20 × ROX Reference Dye (ngati pakufunika) mu bokosi ndi ayezi, kupanga izi kusungunuka mwachibadwa, ndi kusakaniza mofatsa.
B: Konzani dongosolo la RT-qPCR
Pezani theka la voliyumu ya zomwe zimachitika (mwachitsanzo, ngati voliyumu ya makinawo ndi 20 μL, ikufunika kupeza 10 μL 2 × Direct RT-qPCR Buffer). Zakuchuluka kwa enzyme, tikuyembekeza kutiM-MLV: 10-30 U / 20 μL kachitidwe kachitidwe,TaqDNA polymerase:1-2 U / 20 μL reaction systemili ndi lingaliro lathu, muyenera kuyesa ndikusintha), onjezani template ya RNA, zoyambira zenizeni ndi zofufuza, ndikuwonjezera RNase-Free ddH2O mpaka 20 μL.Kukonzekera kwapadera kwa RT-qPCR reaction system kungatchulidwe patebulo lotsatirali 1.
Gulu 1: Konzani dongosolo la RT-qPCR
| Chigawo | Voliyumu | Kukhazikika komaliza |
| 2 × Direct RT-qPCR Buffer | 10 μl pa | 1 × pa |
| Foreasy Reverse Transcriptase(MMLV) | 10-30 U | |
| Foreasy HS Taq DNA Polymerase | 1-2 U | |
| Woyamba Patsogolo (10 μM) | 0.8 μl | 50-900nM |
| Choyambira Chosinthira (10 μM) | 0.8 μl | 50-900nM |
| Kufufuza (4 μM) | 1 ml | 200nM |
| Template (RNA kapena Lysate) | X μL | |
| 20 × ROX Reference Dye | - | 1* |
| RNase-FreeddH2O | (6.4-X) μL | |
| Chiwerengero chonse | 20 μl pa |
Chidziwitso: Forward Primer ndi Reverse Primer ndi zoyambira zenizeni za jini yomwe mukufuna.Dongosolo la qPCR litha kusinthidwa molingana ndi kuyesa kothandiza ndi mitundu ya PCR.Tikupangira 400nM pakuphatikiza komaliza kwa zoyambira zambiri.Sinthani kuchuluka kwa zoyambira zenizeni ndi ma probes molingana ndi ndende yokonzedwa komanso ndende yomaliza yovomerezeka, chonde.
1 *: Sankhani komaliza komaliza kwa ROX Reference Dye malinga ndi zida zosiyanasiyana za PCR.Kuphatikizika koyenera kwa ROX Reference Dye kwa zida zodziwika bwino za PCR kukuwonetsedwa patebulo lotsatirali:
| Zida zambiri za PCR | Magulu omaliza a ROX Reference Dye |
| ABI PRISM 7000/7300/7700/7900HT/Step One,ndi zina. | 1 × (mwachitsanzo, onjezani 1μl 20 × ROX Reference Dye mu 20 μL reaction system) |
| ABI 7500, 7500 Fast, Stratagene Mx3000P, Mx3005P ndi Mx4000, etc. | 0.5× (mwachitsanzo, onjezani 0.5μl 20×ROX Reference Dye mu 20 μL reaction system) |
| Roche PCR chida, Bio-Rad PCR chida, Eppendorf kuchuluka PCR chida, etc. | Popanda ROX Reference Dye |
Kusungirako ndi moyo wa alumali
Zida ziyenera kusungidwa pa -20 ± 5 ℃.Iyenera kusungidwa nthawi yomweyo mu -20 ℃ thermostatic firiji.Nthawi yovomerezeka ndi zaka 2 ngati isungidwa pamalo abwino.
