Mashiinnada Dhagaxa ee Shiinaha
Xirmadani waxay adeegsataa nidaam u gaar ah lysis-dhise kaas oo si dhakhso ah uga sii deyn kara RNA muunadaha unugyada dhaqameed ee falcelinta RT-qPCR, taas oo meesha ka saaraysa habka nadiifinta RNA waqti-qaadka iyo tacabka badan.Qaabka RNA waxa lagu heli karaa 7 daqiiqo oo kaliya.5× Toos ah RT Mix iyo 2× Toos ah qPCR Mix-SYBR reagents ay bixiso qalabku waxay si degdeg ah oo wax ku ool ah u heli karaan natiijooyinka PCR-ga dhabta ah.
5× Toos ah RT Mix iyo 2× qPCR Mix-SYBR toos ah waxay leeyihiin dulqaad adag oo ka hortag ah, iyo lysate ee shaybaarada waxaa loo isticmaali karaa qaab-dhismeedka RT-qPCR si toos ah.Qalabkani waxa uu ka kooban yahay RNA-ga gaarka ah ee xidhiidhka sare ee Foregene reverse transcriptase, iyo Hot D-Taq DNA polymerase, dNTPs, MgCl2, falcelinta falcelinta, PCR optimizer iyo stabilizer.
200×20μl Rxns, 1000×20μl Rxns
Qaybta I | Buffer CL |
Foregene Protease Plus II | |
Buffer ST | |
Qaybta II | Tirtiraha DNA |
5× Toos ah RT Mix | |
2× Toos ah qPCR Mix-SYBR | |
50× ROX Tixraaca Midabka | |
RNase-free ddH2O | |
Tilmaamaha |
■ Fudud oo wax ku ool ah: tignoolajiyada Cell Direct RT, muunadaha RNA waxaa lagu heli karaa 7 daqiiqo oo keliya.
■ Baahida muunada waa yar tahay, maadaama ay yar tahay 10 unug ayaa la tijaabin karaa.
■ Soo saarista sare: waxay si dhakhso ah u ogaan kartaa RNA ee unugyada lagu beeray 384, 96, 24, 12, 6-cells.
■ Tirtiraha DNA-da si dhakhso ah ayuu u saari karaa genome-yada la sii daayay, si weyn ayuu u yareynayaa saameynta natiijooyinka tijaabada dambe.
■ Nidaamka RT iyo qPCR ee la hagaajiyay ayaa ka dhigaya laba-tallaabo dib-u-celinta RT-PCR mid hufan iyo PCR mid gaar ah, oo aad u adkeysi badan ka-hortagga falcelinta RT-qPCR.
Baaxadda codsiga: unugyada dhaqanka.
- RNA lagu sii daayay muunad lysis: kaliya lagu dabaqi karo qaabka RT-qPCR ee xirmadan.
-Qalabka waxa loo isticmaali karaa ujeedooyinka soo socda: falanqaynta muujinta hidde-sidaha, xaqiijinta saamaynta aamusinta hidde-sidaha ee siRNA, baadhista daroogada, iwm.
Qaybta I ee xirmadan waa in lagu kaydiyaa 4℃;Qaybta II waa in lagu kaydiyaa at -20 ℃.
Foregene Protease Plus II waa in lagu kaydiyaa 4 ℃, ha ku barafaysan -20 ℃.
Reagent 2× Toos ah qPCR Mix-SYBR waa in lagu kaydiyaa -20℃ gudcurka;haddii si joogto ah loo isticmaalo, waxa kale oo lagu kaydin karaa 4℃ kaydinta muddada gaaban (la isticmaalo 10 maalmood gudahood).
Marka loo eego baahiyahaaga dhabta ah, dooro habka guud ee naqshadaynta iyo qorshaynta ugu macquulsan