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Ibyerekeye gukuramo RNA

Imyaka 30 irashize, kugirango ikurwemo RNA, icyo twatekerezaga nukuntu twabona RNA yuzuye, ariko, nticyari gikenewe kugirango umuvuduko wo kuvoma (Igisekuru cya mbere Techology uburyo bwa Trizol yavutse).Hamwe no kuvugurura iterabwoba rya RNA ikuramo, ibyo dushobora gukora ntabwo bigarukira gusa kubisubizo.Iyo bahuye nogukuramo ibisekuruza byambere bya Trizol bishingiye kuri reagent, abashakashatsi muri rusange bahangayikishijwe nintambwe zayo zitoroshye;nubwo nyuma yo gutezimbere no guhanga udushya, byafashe igihe kinini.Muri icyo gihe, reagent kama nka fenol na chloroform igomba gukoreshwa mugikorwa cyo kuyikuramo, nayo ibangamira umutekano w abakozi bireba kurwego runaka.

2Bikora neza, byoroshye kandi bifite umutekano

Kubona ibicuruzwa bikuramo neza, byoroshye kandi byizewe byahindutse intego yibanze yabashakashatsi.Kubwibyo, uburyo bwa Silica Spin Inkingi ya adsorption yo gukuramo RNA yabayeho.Ibicuruzwa bya Foregene byo mu gisekuru cya gatatu biva mu bicuruzwa biva mu gisekuru cyabanjirije kuvoma inkingi, ukoresheje (ADN-Cleaning + RNA gusa) ikoranabuhanga rya Spin Column ebyiri kugirango ikureho gDNA hamwe n’imyanda icyarimwe, nta DNase ivura gDNA isigaye, kandi ntigikoreshwa na fenolike, n'ibindi.

Imbonerahamwe yo kugereranya ibikorwa

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Imbonerahamwe yubushakashatsi igereranya imbonerahamwe

Foregene yuzuye ya RNA yo kwigunga ifite umusaruro mwinshi wo gukuramo RNA, ubuziranenge bwiza, kandi ingaruka zo gukuramo miRNA nibyiza kuruta ibindi bikoresho

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Electrophoresis ya 50 mg yumwijima wimbeba RNA yose yakuwe muburyo butandukanye
Igisekuru cya mbere (1: Trizol)
Igisekuru cya kabiri (2: ibikoresho byo gukuramo RNA muri sosiyete A)
Igisekuru cya gatatu (3: ibikoresho byo gukuramo RNA wongeyeho muri sosiyete A, 4: Foregene: ibikoresho bya RNA byo kwigunga)

Igishushanyo cya QPCR (igishushanyo mbonera cya amplification)

ADN ya genomic igomba gukurwaho neza mugihe cyo gukuramo RNA, bitabaye ibyo ikagira ingaruka zitagenzurwa kubushakashatsi bwo hasi (agaciro ka CT kare, ibyiza).Foregene igisekuru kizaza kirashobora gukuraho neza ADN genomic hatabayeho kongeramo DNase, bigatwara umwanya munini, mugihe wirinze kwangirika guterwa na RNase exogenous.
Igisekuru cya mbere (icyatsi cyerekana TRlzol-, umutuku werekana TRlzol +)

5Igisekuru cya kabiri (Ubururu bwerekana Isosiyete ikuramo RNA -, umutuku yerekana Isosiyete A RNA ikuramo ibikoresho +)

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Igisekuru cya gatatu (ubururu bwerekana isosiyete ikuramo ibikoresho bya RNA wongeyeho, icyatsi cyerekana Foregene RNA ibikoresho byo kwigunga)

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Gukuramo RNA byuzuye muri 11min

Ntibikenewe kumyaka yo gukuramo acide nucleic uburambe bwo gukusanya, Foregene-ikora neza cyane hamwe na RNA yo kwigunga ibikoresho byuruhererekane birashobora kugufasha kubona umusaruro mwinshi kandi ufite isuku nyinshi RNA muminota 11 kugirango uhuze ibyifuzo bitandukanye byo munsi yubushakashatsi.

9Kugeza ubu, Foregene yose hamwe RNA isolaton kits yuruhererekane rwibicuruzwa byamenyekanye numubare munini wabakiriya kandi byagize uruhare mubitabo byinshi bitanga amanota menshi.Umukiriya yakoresheje muburyo bwo gutanga ibitekerezo byerekana imbonerahamwe ya electrophoresis hamwe numubare wuzuye wa PCR amplification curve nyuma yo kwandukura inyandiko zerekana ko RNA yakuwe mugukoresha ibicuruzwa bya Foregene itari hejuru cyane mubitekerezo, ahubwo no mubunyangamugayo.Gusa tekinoroji yibicuruzwa yageragejwe nisoko irashobora gutoneshwa nabantu bose.

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Akagari Igiteranyo cya RNA Igikoresho cyo kwigunga

Umusaruro mwinshi wa RNA

Byihuse: Kurangiza RNA kwigunga muminota 11

Umutekano: Nta miti kama yongeyeho

Amwe mumagambo menshi yatanzwe:

1. Yuan Fang, Zezhong Liu, Yang Qiu, n'abandi.Kubuza virusi ya poroteyine ya RNAi na peptide yabashushanyije irinda kwandura virusi muri vivo, Ubudahangarwa, 54 (10), 2021: 2231-2244.e6.(NIBA: 31.745,Virusi RNA Igikoresho cyo kwigunga, Injangwe No.RE-02011)

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

2. Ren, Y., Wang, A., Wu, D. n'abandi.Kubuza ubudahangarwa bw'umubiri hamwe na apoptose na poroteyine ya cytomegalovirus ya muntu UL37x1 ituma virusi ikwirakwizwa neza.Nat Microbiol 7, 1041–1053 (2022).(NIBA: 30.964,Akagari Igiteranyo cya RNA Igikoresho cyo kwigunga, Injangwe No.RE-03111)

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


Igihe cyo kohereza: Nyakanga-27-2022