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Nhanganyaya:

RNA, zita resainzi ribonucleic acid, chidimbu chayo chinobva kuRiboNucleic Acid, imhando ye nucleic acid inoumbwa nekubatanidza angangoita gumi nemaviri eribonucleotides kuburikidza nephosphodiester bond.

Mhedziso yekuongorora RNA ndeyekubvisa kusvibiswa-isina Total RNA, uye musimboti wekutanga weRNA kubviswa unogona kupfupikiswa sezvizvi:

1. Kuvhiringidza masero kutanga

2. Wobvisa macromolecular tsvina (mapuroteni, lipids, DNA, nezvimwewo)

3. Pakupedzisira, iyo RNA inonaya ichibvisa munyu nezvinonyungudutsa, uye ichiwedzera kuchenesa RNA.

Yakapfuura uye yazvino nzira yekubvisa

The first generation of RNA extraction method is TRIzol precipitation method (mareagents anoshandiswa anosanganisira guanidine isothiocyanate, phenol, chloroform, etc.) yakagadzirwa nasekuru vanonzi Piotr Chomczynski nambuya Nicoletta Sacchi, nokuti chigadzirwa ichi chinobvumira RNA kubudiswa Kurumidza uye nyore, kubva panguva iyo yakatanga RNA tsvakurudzo.

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Gare gare, Sekuru vakatengesa patent iyi kuInvitrogen, uye Invitrogen yakasimudzira nzira iyi kupasi rose ikava nzira inoshandiswa nevakawanda.

Gare gare, Invitrogen yakawanikwa neThermo!

Chigadzirwa chinomiririra: Invitrogen's TRIzol reagent

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Nekudaro, iyo TRIzol kunaya nzira inoda kunaya kwakawanda uye kubviswa kweRNA, uye mareagents anoshandiswa ane muchetura uye anotora nguva yakareba kushanda.

Naizvozvo, chizvarwa chechipiri chekororamu-enzyme yekuchenesa nzira yakavapo, ichishandisa silica gel membrane seyakanyanya nucleic acid adsorption zvinhu kuti iwedzere kudzoreredza kweRNA mumuenzaniso uchibvisa kumwe kusvibiswa.

Nzira yechizvarwa chechipiri inorerutsa kushanda pahwaro hwekushanda kwakagadzikana, inoderedza njodzi, uye inovandudza kushanda kweRNA kutorwa.

Zvigadzirwa zvinomiririra: QIAGEN RNeasy uye yakawanda zvigadzirwa zvepamba

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Nekudaro, nzira dzese dzekutanga neyechipiri dzechizvarwa dzinoda kushandiswa kweDNase kubvisa kusvibiswa kweDNA, izvo zvinounza kumwe kusagadzikana kune kunotevera kuyedza kuzasi.Residual DNase inoderedza kugona kwe reverse transcription uye kuderedza yekutanga-strand cDNA synthesis, zvichizokanganisa kurongeka kwemhedzisiro.

Nokudaro, chizvarwa chechitatu chekuchera nzira inogadziriswa pamusana pechizvarwa chechipiri.Kuedza hakudi kuwedzera chero ma enzymes, uye RNA inogona kuwanikwa nyore nemakoramu maviri chete.

Column 1: DNA-Cleaning Column kujekesa DNA

Column 2: RNA-chete Column inonyanya adsorbs uye inodzosera RNA

Kushanda kwakapfava kwakadaro hakungochengete nguva chete, asiwo kunofambisa kushanda kwezviyedzo zvepasi!

Zvigadzirwa zvinomiririra: QIAGEN RNeasy Plus uyeForegeneRNA yekuchenesa yakatevedzana zvigadzirwa

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(RNeasy Plus)

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(Forgene'sPlant Yese RNA Isolation Kit)

Taura iwe chinyararire!Foregene's RNA kit zvakare inopa yakanyanya nyore kushandisa RNase Eraser, inogona kubvisa zvachose 400μg yeRNAse panzvimbo yakasimba muminiti 1, pasina kuwedzera chero chepfu kumuviri wemunhu!

Pic1: Operational process kuenzanisa chati

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Zvigadzirwa zveForegene zvinogona kupedzisa RNA kutorwa munziyo mbiri chete, usaite vamwe godo nekufambira mberi kwekuedza kwako!

Pic2:Experimental results kuenzanisa chati

Chizvarwa chekutanga (1: TRIzol)

Chizvarwa chechipiri (2: RNeasy of QIAGEN)

Chizvarwa chechitatu (3: QIAGEN's RNeasy plus, 4: Foregene)

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Pic3: qPCR girafu (amplification curve girafu)

Chizvarwa chekutanga (girinhi zvinoreva TRIzol -, tsvuku zvinoreva TRIzol +)

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Chizvarwa chechipiri (denga rebhuruu rinoreva QIAGEN RNeasy-, tsvuku zvinoreva QIAGEN RNeasy +)

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Chizvarwa chechitatu (bhuruu zvinoreva QIAGEN RNeasy plus, green zvinoreva Foregene)

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(Cherechedza: qPCR graph "-" inoratidza kuti DNase haisati yawedzerwa uye kusvibiswa kweDNA hakuna kubviswa. Nokudaro, CT yakakwirira uye mhedzisiro yacho haina kururama; "+" inoratidza kuti DNase yakawedzerwa uye kusvibiswa kweDNA kwabviswa, chigumisiro chakarurama)

Yese yekuvandudza tekinoroji inoreva kuti kushanda kwekuyedza kuri kusimukira mune yakapfava uye yakawanda-mushandisi-ane hushamwari gwara.

Foregene ine chinangwa chekubatsira vaongorori vesainzi kuchengetedza nguva yekuyedza uye mutengo, kuwana mhedzisiro yemhando yepamusoro yekuyedza, uye pamwe chete kugadzira zvakakosha zvakawanikwa patsvagiridzo yesainzi.


Nguva yekutumira: Jul-09-2021