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Malunga nokutsalwa kwe-RNA

Kwiminyaka engamashumi amathathu eyadlulayo, ekutsalweni kwe-RNA, into ebesinokuyicinga kuphela indlela yokufumana i-RNA epheleleyo, nangona kunjalo, kwakungekho mfuneko yesantya sokutsalwa (Isizukulwana sokuqala seTekhnoloji indlela yeTrizol eyazalwa).Ngohlaziyo oluphindaphindiweyo lwetekhnoloji yokutsalwa kwe-RNA, into esinokuyenza ayiphelelanga ekufumaneni iziphumo.Xa ujongene nokutsalwa kwee-reagents ezisekelwe kwisizukulwana sokuqala se-Trizol, abenzi bovavanyo babexhatshazwa ngokubanzi ngamanyathelo ayo okusebenza anzima;nangona emva kokulungiswa okuninzi kunye nezinto ezintsha, kuthathe ixesha elininzi.Ngexesha elifanayo, i-reagents ye-organic efana ne-phenol kunye ne-chloroform kufuneka isetyenziswe kwinkqubo yokukhutshwa, ekwasongela ukhuseleko lwabasebenzi abachaphazelekayo kwinqanaba elithile.

2Isebenza kakuhle, iluncedo kwaye ikhuselekile

Ukufumana imveliso yokukhutshwa kwe-RNA esebenzayo, efanelekileyo kwaye ekhuselekileyo ibe yinjongo ephambili yabavavanyi.Ke ngoko, indlela ye-Silica Spin Column adsorption yokutsalwa kwe-RNA ibekho.Imveliso ye-Foregene yesizukulwana sesithathu sokutsalwa kwe-RNA zoba kwiinzuzo zemveliso yokukhutshwa kwekholamu yesizukulwana sangaphambili, usebenzisa (i-DNA-Cleaning + RNA kuphela) iteknoloji ezimbini ze-Spin Column ukususa i-GDNA kunye ne-debris ye-tissue ngexesha elifanayo, ngaphandle kwe-DNase unyango lwe-gDNA eseleyo, kwaye ayisayi kusebenzisa i-phenol, njl.

Itshathi yokuthelekisa inkqubo yokusebenza

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Itshathi yokuthelekisa iziphumo zovavanyo

Ikhithi yokuhlukaniswa ye-RNA ye-Foregene inemveliso ephezulu yokutsalwa kwe-RNA, ukucoceka okuhle, kunye nesiphumo sokutsalwa kwe-miRNA singcono kunezinye iikhithi zophawu.

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I-Electrophoresis ye-50 mg yesibindi semouse iyonke i-RNA ekhutshwe ngeendlela ezahlukeneyo
Isizukulwana sokuqala (1: Trizol)
Isizukulwana sesibini (2: Ikhithi yokutsalwa kwe-RNA kwinkampani A)
Isizukulwana sesithathu (3: Ikhithi yokutsalwa kwe-RNA kunye nenkampani A, 4: Foregene:RNA Isolation kits)

Igrafu ye-qPCR (igrafu yokukhulisa igophe)

I-DNA ye-Genomic kufuneka isuswe ngokufanelekileyo ngexesha lokukhutshwa kwe-RNA, ngaphandle koko kuya kuba nemiphumo engalawulekiyo kwiimvavanyo ezisezantsi (ixabiso le-CT lokuqala, iimpawu zobuxoki).Ikhithi yesizukulwana esilandelayo inokususa ngokugqibeleleyo i-DNA ye-genomic ngaphandle kokongezwa kwe-DNase, igcina ixesha elininzi, ngelixa ithintela ukuthotywa okubangelwa kungcoliseko lwe-RNase yangaphandle.
Isizukulwana sokuqala (uhlaza lubonisa i-TRlzol-, ebomvu ibonisa i-TRlzol +)

5Isizukulwana sesibini (I-Blue ibonisa iNkampani Ikhithi yokutsalwa kwe-RNA -, ebomvu ibonisa iNkampani Ikhithi yokutsalwa kwe-RNA +)

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Isizukulwana sesithathu (eblue ibonisa inkampani A RNA extraction kit plus, green ibonisa Foregene RNA Isolation kits)

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(Qaphela: "_" kwigrafu ye-qPCR ibonisa ukuba i-DNase ayidityaniswanga, kwaye ukungcoliswa kwe-DNA akukasuswa; "+" ibonisa ukuba i-DNase yongeziwe, kwaye ukungcoliswa kwe-DNA kususiwe)

Gqibezela ukukhutshwa kwe-RNA ngaphakathi kwe-11min

Akukho mfuneko yeminyaka yokuqokelelwa kwamava okutsalwa kwe-nucleic acid, i-Foregene ephezulu esebenzayo iyonke i-RNA yokwahlula iikhithi zothotho lweemveliso zinokukuvumela ukuba ufumane isivuno esikhulu kunye nokucoceka okuphezulu kwe-RNA ngaphakathi kwemizuzu eli-11 ukuhlangabezana neemfuno ezahlukeneyo zokulinga.

9Ukuza kuthi ga ngoku, i-Foregene iyonke ye-RNA isolaton kits iimveliso ziye zaqatshelwa linani elikhulu labathengi kwaye zibe negalelo kuncwadi oluninzi olunamanqaku aphezulu.Ukusetyenziswa kwangempela komthengi kwitshathi ye-electrophoresis yempendulo kunye ne-quantitative PCR amplification curve emva kwe-reverse transcription ibonisa ukuba i-RNA ekhutshwe ngokusetyenziswa kweemveliso ze-Foregene ayiphezulu kuphela ekugxininiseni, kodwa kunye nokuthembeka okulungileyo.Kuphela iteknoloji yemveliso eye yavavanywa yimarike inokuthandwa ngumntu wonke.

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ISeli iyonke yeRNA Isolation Kit

Isivuno esiphezulu se-RNA

Ukukhawuleza: Gqiba i-RNA Isolation kwimizuzu eli-11

Ukhuseleko: Akukho khemikhali ye-Organic eyongeziweyo

Ezinye izicatshulwa zamanqaku aphezulu:

1. Yuan Fang, Zezhong Liu, Yang Qiu, et al.Ukuvinjelwa kwe-viral suppressor ye-RNAi iiprotheni nge-designer peptides ikhusela ukusuleleka kwi-enteroviral kwi-vivo, i-Immunity, i-54 (10), i-2021: 2231-2244.e6.(Ukuba:31.745,IViral RNA Isolation Kit,Inombolo yekati.RE-02011)

https://www.sciencedirect.com/science/article/pii/S1074761321003617

2. URen, Y., Wang, A., Wu, D. et al.Ukuvinjelwa kabini kwe-innate immunity kunye ne-apoptosis yiprotheni ye-cytomegalovirus yabantu i-UL37x1 yenza ukuphindaphinda kwentsholongwane okusebenzayo.I-Nat Microbiol 7, 1041-1053 (2022).(Ukuba:30.964,ISeli iyonke yeRNA Isolation Kit,Inombolo yekati.RE-03111)

https://doi.org/10.1038/s41564-022-01136-6


Ixesha lokuposa: Jul-27-2022