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ISeli Ngqo RT qPCR Kit—SYBR GREEN I Direct Cell Lysis Cell Ready Inyathelo elinye qRT-PCR Kits

Inkcazelo yeKit:

Ikati.No.DRT-01011/01012

Kwiseli ngqo RT-qPCR usebenzisa 10-1,000,000 iiseli,

kunye nokwenza i-RT-qPCR ngqo kwiiseli ngaphandle kokucocwa kwangaphambili kwe-RNA.

Iiseli ze-lysed ngokuthe ngqo ukukhulula i-RNA ye-RT-qPCR;inkqubo yonyamezelo ephezulu yenza kube yimfuneko ukuhlanjululwa kwe-RNA kunye nokusebenzisa ngokuthe ngqo i-lysates yeseli njengeetemplates ze-RNA zokusabela kwe-RT.Ukukhawuleza kwaye kulula;uvakalelo oluphezulu, ukuchaneka okuqinileyo, kunye nokuzinza okulungileyo.

◮Ilula kwaye iyasebenza: ngeTekhnoloji ye-Cell Direct RT, iisampulu ze-RNA zinokufumaneka kwimizuzu eyi-7 nje.

Imfuno yesampulu incinci, iphantsi njengeeseli ezili-10 zinokuvavanywa.

◮Ukuphumelela okuphezulu: inokubona ngokukhawuleza i-RNA kwiiseli ezikhuliswe kwii-384, 96, 24, 12, 6-well plates.

I-DNA Eraser inokususa ngokukhawuleza i-genomes ekhutshiweyo, inciphise kakhulu impembelelo kwiziphumo zovavanyo ezilandelayo.

Inkqubo ye-RT elungiselelweyo kunye ne-qPCR yenza amanyathelo amabini e-RT-PCR aguqulelwe umva asebenze ngakumbi kwaye i-PCR icace ngakumbi, kwaye ixhathisa ngakumbi kwi-RT-qPCR reaction inhibitors.


Iinkcukacha zeMveliso

Iithegi zeMveliso

FAQ

Khuphela izixhobo

Iinkcazelo

Le khithi isebenzisa inkqubo ye-lysis buffer ekhethekileyo enokuthi ikhulule ngokukhawuleza i-RNA kwiisampulu zeeseli ezikhuliswe kwi-RT-qPCR reactions, ngaloo ndlela isusa inkqubo yokucoca i-RNA ethatha ixesha kunye nenzima.Ithemplethi ye-RNA inokufumaneka ngemizuzu nje esi-7.I-5×Ngqo RT Mix kunye 2×Iirejenti ezithe ngqo ze-qPCR Mix-SYBR ezibonelelwe yikiti zinokukhawuleza nangempumelelo ukufumana iziphumo zePCR zexesha lokwenyani.

5×Ngqo RT Mix kunye 2×Ngokuthe ngqo i-qPCR Mix-SYBR inokunyamezela inhibitor eqinile, kwaye i-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 iyasebenza : ngeTekhnoloji ye-Cell Direct RT, iisampuli ze-RNA zinokufumaneka kwimizuzu eyi-7 nje.

∎ 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

Iseli ethe ngqo kwidayagram ye-RT qPCR

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 ngo-4℃, musa ukuba ngumkhenkce ku -20℃.

 I-Reagent 2×Direct qPCR Mix-SYBR kufuneka igcinwe -20ebumnyameni;ukuba isetyenziswa rhoqo, inokugcinwa ngo-4℃ kugcino lwexesha elifutshane (sebenzisa ngaphakathi kweentsuku ezili-10).


  • Ngaphambili:
  • Okulandelayo:

  • 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


    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


    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

     

    3.qPCR Mix


    Umxube we-qPCR

    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

    Izikhokelo zoMyalelo:

    I-QuickEasyTM Cell Direct RT-qPCR Kit-SYBR Green I

    Bhala umyalezo wakho apha kwaye uwuthumele kuthi