Umthombo wefektri iChina Rt PCR Assay Reagent ye-2019 New Virus (ORF1ab, N gene)
Thatha uxanduva olupheleleyo lokwanelisa zonke iimfuno zabathengi bethu;ukufikelela kwinkqubela phambili ngokuthengisa uphuhliso lwabathengi bethu;khula ube liqabane lokugqibela lentsebenziswano elisisigxina labathengi kwaye songeze umdla wabathengi kumthombo weFactory I-China Rt PCR Assay Reagent ye-Virus eNtsha ka-2019 (ORF1ab, N gene), Siyaziqinisekisa ukuba siyayila impumelelo emangalisayo ngelixa sisenakho.Besizingela ukuba ngomnye wababoneleli bakho abathembekileyo.
Thatha uxanduva olupheleleyo lokwanelisa zonke iimfuno zabathengi bethu;ukufikelela kwinkqubela phambili ngokuthengisa uphuhliso lwabathengi bethu;khula ube liqabane lokugqibela lentsebenziswano esisigxina yabathengi kwaye ukwandise umdla wabathengiChina PCR Kit, Ikhithi yokuFumana iPcr, Sijonge phambili, siya kugcina isantya kunye namaxesha, siqhubeka nokudala iimveliso ezintsha.Ngeqela lethu lophando olomeleleyo, amaziko emveliso ephucukileyo, ulawulo lwezenzululwazi kunye neenkonzo eziphezulu, siya kubonelela ngeemveliso ezikumgangatho ophezulu kubathengi bethu kwihlabathi liphela.Sikumema ngokunyanisekileyo ukuba ube ngamahlakani ethu oshishino ukuze ufumane izibonelelo ezifanayo.
Iinkcazelo
Le khithi isebenzisa inkqubo ye-lysis buffer ekhethekileyo enokuthi ikhulule ngokukhawuleza i-RNA kwiisampulu zeseli ezikhuliswe kwiimpendulo ze-RT-qPCR, ngaloo ndlela iphelisa inkqubo yokucoca i-RNA ethatha ixesha kunye nenzima.Ithemplethi ye-RNA inokufumaneka kwimizuzu nje esi-7.Umxube we-RT oyi-5 × ngokuthe ngqo kunye ne-2 × Direct qPCR Mix-SYBR ii-reagents ezibonelelwe yikhithi zinokufumana ngokukhawuleza nangempumelelo iziphumo zePCR zexesha lokwenyani.
I-5 × Direct RT Mix kunye ne-2 × Direct qPCR Mix-SYBR inokunyamezela inhibitor eqinile, kunye ne-lysate yeesampuli zingasetyenziswa njenge template ye-RT-qPCR ngokuthe ngqo.Le khithi iqulethe i-RNA ephezulu ehambelana ne-Foregene reverse transcriptase, kunye ne-Hot D-Taq DNA polymerase, i-dNTPs, i-MgCl2, i-reaction buffer, i-PCR optimizer kunye ne-stabilizer.
Iinkcukacha
200×20μl Rxns, 1000×20μl Rxns
Amacandelo ekhithi
Icandelo I | Isithinteli CL |
Foregene Protease Plus II | |
Isithinteli ST | |
Icandelo II | DNA Eraser |
5 × Ngqo RT Mix | |
2 × Ngqo qPCR Mix-SYBR | |
I-50 × i-ROX Reference Dye | |
RNase-Free ddH2O | |
Imiyalelo |
Iimpawu&izinto eziluncedo
■ Ilula kwaye isebenzayo : ngeteknoloji ye-Cell Direct RT, iisampuli ze-RNA zinokufumaneka kwimizuzu nje eyi-7.
∎ Imfuno yesampulu incinci, ingaphantsi njengeeseli ezili-10 zinokuvavanywa.
■ Ukugqithisa okuphezulu: inokubona ngokukhawuleza i-RNA kwiiseli ezikhuliswe kwii-384, 96, 24, 12, 6-well plates.
■ I-DNA Eraser inokususa ngokukhawuleza iigenomes ezikhutshiweyo, inciphise kakhulu impembelelo kwiziphumo zovavanyo ezilandelayo.
■ Inkqubo elungiselelwe i-RT kunye ne-qPCR yenza amanyathelo amabini e-RT-PCR akhutshelwe umva asebenze ngakumbi kwaye i-PCR icace ngakumbi, kwaye imelane ngakumbi ne-RT-qPCR reaction inhibitors.
Isicelo seKit
Ububanzi besicelo: iiseli ezikhuliswe.
- I-RNA ekhutshwe ngesampuli ye-lysis: isebenza kuphela kwi-template ye-RT-qPCR yale khithi.
- Ikiti ingasetyenziselwa ezi njongo zilandelayo: uhlalutyo lwe-gene expression, ukuqinisekiswa kwe-siRNA-mediated gene silence effect, ukuhlolwa kweziyobisi, njl.
Umzobo
Ugcino kunye nobomi beShelf
Inxalenye I yale khithi kufuneka igcinwe kwi-4℃;Icandelo II kufuneka ligcinwe kwi -20 ℃.
I-Foregene Protease Plus II kufuneka igcinwe ku-4℃, ungakhenkcezisi ku -20℃.
I-Reagent 2 × Direct qPCR Mix-SYBR kufuneka igcinwe kwi -20 ℃ ebumnyameni;ukuba isetyenziswe rhoqo, inokugcinwa kwi-4 ℃ ukwenzela ukugcinwa kwexesha elifutshane (ukusetyenziswa ngaphakathi kweentsuku ezili-10). Thatha umsebenzi opheleleyo ukwanelisa zonke iimfuno zabaxhasi bethu;ukufikelela kwinkqubela phambili ngokuthengisa uphuhliso lwabathengi bethu;khula ube liqabane lokugqibela lentsebenziswano elisisigxina labathengi kwaye songeze umdla wabathengi kumthombo weFactory I-China Rt PCR Assay Reagent ye-Virus eNtsha ka-2019 (ORF1ab, N gene), Siyaziqinisekisa ukuba siyayila impumelelo emangalisayo ngelixa sisenakho.Besizingela ukuba ngomnye wababoneleli bakho abathembekileyo.
Umthombo wefektriChina PCR Kit, I-PCR Detection Kit, Sijonge phambili, siya kugcina isantya kunye namaxesha, siqhubeka nokudala iimveliso ezintsha.Ngeqela lethu lophando olomeleleyo, amaziko emveliso ephucukileyo, ulawulo lwezenzululwazi kunye neenkonzo eziphezulu, siya kubonelela ngeemveliso ezikumgangatho ophezulu kubathengi bethu kwihlabathi liphela.Sikumema ngokunyanisekileyo ukuba ube ngamahlakani ethu oshishino ukuze ufumane izibonelelo ezifanayo.
Ixesha lokwenyani imigaqo yoyilo lwe-PCR primer
I-Primer yokuPhambili kunye ne-Primer yokuBuyisa
Ngexesha lokwenyani iPCR, uyilo lweprimer lubaluleke kakhulu.Iiprimers zinxulumene nobucacileyo kunye nokusebenza kakuhle kokwandiswa kwePCR, kwaye zinokuyilwa ngokubhekiselele kule migaqo ilandelayo:
Ubude bePrimer: 18-30bp.
Umxholo weGC: 40-60%.
Ixabiso le-Tm: I-software yoyilo lwe-Primer, njenge-Primer 5, inokunika ixabiso le-Tm le-primer.Amaxabiso e-Tm ee-primers ezinyukayo nezisezantsi kufuneka zibe kufutshane kangangoko.Ifomula yokubala ye-Tm nayo ingasetyenziswa: Tm = 4 °C (G + C) + 2 °C (A + T).Xa kusenziwa i-PCR, iqondo lobushushu elingaphantsi kwexabiso le-primer Tm le-5 °C likhethwa ngokubanzi njengeqondo lobushushu lokutshisa (ukunyuka okuhambelanayo kwiqondo lobushushu lobushushu kunokunyusa ukuchana kwe-PCR reaction).
Iiprimers kunye neemveliso zePCR:
I-design primer PCR yokukhulisa ubude bemveliso ikhethwa yi-100-150bp.
Ii-primers zoyilo kwindawo yesibini yesakhiwo setemplate kufuneka igwenywe kangangoko kunokwenzeka.
Gwema ukubunjwa kweziseko ezi-2 okanye ngaphezulu ezihambelanayo phakathi kwee-3′ zeziphelo ze-primers ezinyukayo kunye nezantsi.
Isiseko se-Primer 3′ asikwazi ukubakho nge-3 eyongezelelweyo elandelelanayo i-G okanye i-C.
Iiprimers ngokwazo azikwazi ukuba nezakhiwo ezincedisayo, ngaphandle koko kuya kwenziwa isakhiwo se-hairpin, esichaphazela ukukhulisa i-PCR.
I-ATCG kufuneka isasazwe ngokulinganayo ngokulandelelana kwe-primer, kwaye isiseko se-terminal ye-3 kufuneka sigwenywe njenge-T.
ISihlomelo 1: Ipakethi yecandelo leCell Direct RT-qPCR Kit
1.Iseli Lysis Solution
| |||
Amacandelo ekhithi (Inkqubo ye-lysis ye-24 / kakuhle) | DRT-01011-A1 | DRT-01011-A2 | |
100 T | 500 T | ||
InxalenyeI | Isithinteli CL | 20 ml | 100 ml |
Foregene Protease Plus II | 400 μl | 1 ml × 2 | |
Isithinteli ST | 1 ml × 2 | 10 ml | |
InxalenyeII | DNA Eraser | 400 μl | 1 ml × 2 |
2.RT Umxube
| |
Amacandelo ekhithi (20 μl inkqubo yokusabela) | DRT-01011-B1 |
200 T | |
5 × Ngqo RT Mix | 800 μl |
RNase-Free ddH2O | 1.7 ml × 2 |
| ||
Amacandelo ekhithi (20 μl inkqubo yokusabela) | DRT-01011-C1 | DRT-01011-C2 |
200 T | 1000 T | |
2 × Ngqo qPCR Mix-SYBR | 1 ml × 2 | 1.7 ml × 6 |
I-50 × i-ROX Reference Dye | 40 μl | 200 μl |
RNase-Free ddH2O | 1.7 ml | 10 ml |