Gutera Igiteranyo cya RNA Yigunze Yongeyeho RNA Igikoresho cya RNA Purificaiton kubihingwa bikungahaye kuri Polysaccharide na Polifenol
Ibisobanuro:
Injangwe.No.RE-05021 / 05022/05024
Kugirango usukure RNA yose uhereye kuburugero rusange rwibimera birimo polysaccharide nyinshi hamwe nibice bya polifenol.
Kuramo vuba vuba ubuziranenge bwuzuye RNA mubitegererezo byibimera birimo polysaccharide na polifenol.
RNase-Yubusa Ukoresheje ADN-Isukura Inkingi
Byoroshye - ibikorwa byose birangirira ku bushyuhe bwicyumba
Byihuse - ibikorwa birashobora kurangira muminota 30
Umutekano - nta reagent ikoreshwa 
Ibisobanuro
50 Imyiteguro, 200 Imyiteguro
Igikoresho gikoresha spin inkingi hamwe na formula yakozwe na Foregene, ishobora gukuramo neza-isuku nini kandi yuzuye ya RNA yuzuye mubice bitandukanye byibimera bifite polysaccharide cyangwa polifenol nyinshi.Itanga ADN-Isukura inkingi ishobora gukuraho byoroshye ADN ya genomic muri lysate ndengakamere na tissue.Inkingi ya RNA yonyine irashobora guhuza RNA neza.Igikoresho gishobora gutunganya umubare munini wicyitegererezo icyarimwe.
Sisitemu yose ntabwo irimo RNase, bityo RNA yatunganijwe ntizitesha agaciro.Buffer PRW1 na Buffer PRW2 birashobora kwemeza ko RNA yabonye itanduye na poroteyine, ADN, ion, hamwe n’ibinyabuzima.
Ibigize ibikoresho
| Buffer PSL1, Buffer PS, Buffer PSL2 |
| Buffer PRW1, Buffer PRW2 |
| Ubusa ddH2O, Inkingi-yoza |
| RNA-Inkingi gusa |
Ibiranga ibyiza
Gukorera mubushyuhe bwicyumba (15-25 ℃) mubikorwa byose, nta bwogero bwa barafu hamwe na centrifugation yubushyuhe buke.
Kit Ibikoresho byuzuye RNase-Ubuntu, nta mpamvu yo guhangayikishwa no kwangirika kwa RNA.
■ By'umwihariko bikwiranye no kweza RNA mu ngero z'ibimera bya polysaccharide na polifenol.
Inkingi ya ADN-Isukura Inkingi ihuza cyane na ADN, kugirango ibikoresho bishobore gukuraho ADN yanduye utongeyeho DNase.
Umusaruro mwinshi wa RNA: RNA-Inkingi gusa hamwe na formula idasanzwe irashobora kweza neza RNA.
Speed Umuvuduko wihuse: byoroshye gukora kandi birashobora kurangira muminota 30.
Umutekano: nta reagent ngenga isabwa.
Quality Ubwiza buhanitse: Ibice bya RNA bisukuye bifite isuku nyinshi, bitarimo poroteyine n’ibindi byanduye, kandi birashobora guhura nuburyo butandukanye bwo kugerageza.
Ibipimo byibicuruzwa
Applices Porogaramu yo hasi: synthesis ya mbere ya cDNA, RT-PCR, clone ya molekile, Blot y'Amajyaruguru, nibindi.
Ample Icyitegererezo: Ibimera bishya cyangwa bikonje bya polysaccharide na polifenol
Ingano: 50mg tissue yibihingwa
Ubushobozi ntarengwa bwa RNA bwo guhuza inkingi yo kweza: 80 μg
Volume Ingano yo gukuraho: 50-200 μl
Porogaramu
Birakwiriye gukuramo no kweza RNA yose uhereye kumyumbati y'ibiti bishya cyangwa bikonje (cyane cyane ibibabi byibabi bishya) bifite polysaccharide nyinshi hamwe na polifenol.
Urujya n'uruza rw'akazi
Igishushanyo
Gutera Igiteranyo cya RNA Isolation Kit Plus yatunganije 50mg yamababi mashya ya polysaccharide na polifenol, naho 5% RNA yatunganijwe yapimwe na electrophorei.
1: Umuneke
2: Ginkgo
3: Impamba
4: Amakomamanga
Ububiko hamwe nubuzima bwa Shelf
Iki gikoresho gishobora kubikwa amezi 24 mugihe cyumye mubushyuhe bwicyumba (15-25 ℃);niba ikeneye kubikwa igihe kirekire, irashobora kubikwa muri 2-8 ℃.
Buffer PSL1 irashobora gushirwa kuri 4 ℃ ukwezi 1 nyuma yo kongeramo β-mercaptoethanol (birasabwa kongeramo mugihe kimwe cyubushakashatsi).
Inkingi yacometse
Inkingi imaze gucomeka, umusaruro wa RNA uragabanuka cyangwa ntibishoboka koza RNA, kandi misa ya RNA yabonetse ni mike.
Isesengura rusange:
1. Ikiruhuko ntangarugero ntabwo cyuzuye.
Icyitegererezo cyo kumeneka ntabwo gitera rwose ADN-CLEANING COLUMN guhagarikwa, mugihe bigira ingaruka kumusaruro wa RNA nubwiza.Turasaba inama yo gusya byihuse muri azote ihagije mugihe wamennye ingero, Gerageza kumenagura urukuta rwicyitegererezo, membrane selile nizindi ngingo.Kubitegererezo byibihingwa bya polyol polysaccharide, turagusaba ko ukoresha Ibimera byose RNA ISOLATION KIT PLUS.
2. Iyo unywa icyitegererezo cyindengakamere cyatandukanijwe hamwe na ADN-Isukura Inkingi, imvura ishobora kuba igabanijwe ishobora guhumeka.
Ingirabuzimafatizo zacitsemo ibice zafashwe zizatera inkingi ya RNA-GUSA izahagarikwa mugihe ibikorwa bya RNA adsorption bikorwa (reba intambwe ya 6).Turagusaba ubwitonzi mugihe unywa iyi ndengakamere kugirango wirinde imyanda ya selile.
3. Icyitegererezo umubare wambere ni mwinshi.
Gukoresha urugero rwinshi bizavamo gutandukanya ibyitegererezo bituzuye cyangwa lysis selile ituzuye na Buffer PSL1, bikaviramo guhagarika inkingi yo kweza mugihe cyo kwezwa.Gutera Igiteranyo Cyuzuye RNA Kwigunga Buri cyitegererezo cyogukora ni 50 mg.Kubitegererezo byibihingwa bya polyol polysaccharide, turagusaba ko wagerageza Gutera Igiteranyo Cyuzuye RNA ISOLATION KIT PLUS.
4. Ubushyuhe bwa centrifuge buri hasi cyane.
Inzira yose ya RNA yo kwigunga no kwezwa ikorwa mubushyuhe bwicyumba (20-25°C), usibye ko ingero z'icyitegererezo zavunitse na azote yuzuye. Ubushyuhe bwa centrifuges zimwe na zimwe za kirogenike ziri munsi ya 20℃, zishobora gutera guhagarika ADN-Isukura Inkingi na / cyangwa RNA-Inkingi gusa.Niba ibi bibaye, shyira ubushyuhe bwa centrifuge kuri 20-25℃, namenya neza ko lysis ivanze na / cyangwa etanol yongeyeho supernatant yashushe kuri 37°C.
Nta RNA yakuweho cyangwa umusaruro wa RNA uri muke
Mubisanzwe hariho ibintu byinshi bigira ingaruka kumikorere yo kugarura, nka: icyitegererezo cya RNA, uburyo bwo gukora, ingano ya elution, nibindi.
Isesengura ry'impamvu zisanzwe nkuko bikurikira:
1.Ubwogero bwa ice cyangwa ubushyuhe buke (4 ° C) centrifugation yakozwe mugihe cyo kubaga.
Igitekerezo: Kora ku bushyuhe bwicyumba (15-25°C) mubikorwa byose, ntukore ubwogero bwa ice hamwe na centrifugation yubushyuhe buke.
2. RNA yateshejwe agaciro kubera kubika neza icyitegererezo cyangwa kubika igihe kirekire icyitegererezo.
Icyifuzo: Ingero zegeranijwe vuba zigomba gukonjeshwa vuba muri azote yuzuye, hanyuma zikabikwa kuri -80 ° C igihe kirekire, ukirinda gukonjesha no gukonjesha ingero;cyangwa uhite ushiramo ibyitegererezo muri RNA stabilisateur RNAlater igisubizo (icyitegererezo cyinyamaswa).
3.Ibice bidahagije byo gutandukanya na lysis biganisha ku guhagarika inkingi yo kwezwa.
Igitekerezo: Mugihe cyo gusya ingirangingo, nyamuneka urebe neza ko tissue iri hasi bihagije, kandi uyimure vuba muri Buffer PSL1 yateguwe mbere (wemeze ko igipimo gikwiye cya β-ME cyongeweho, reba intambwe ya 1 yuburyo).
4.Umuhanga wongeyeho nabi.
Igitekerezo: Menya neza ko RNase-idafite ddH2O yatonywe hagati yinkingi yo kweza.
5.Ubunini bukwiye bwa Ethanol yuzuye ntabwo yongewe kuri Buffer PSL2 cyangwa Buffer PRW2.
Igitekerezo: Nyamuneka kurikiza amabwiriza, ongeramo ingano yukuri ya Ethanol yuzuye kuri Buffer PSL2 na Buffer PRW2 hanyuma uvange neza mbere yuko ibikoresho bikoreshwa.
6. Ingano yicyitegererezo ntikwiye.
Igitekerezo: Koresha mg 50 za tissue kuri 500 μl ya Buffer PSL1.Gukoresha tissue nyinshi bizagabanya ingano ya RNA yakuweho kandi isuku ya RNA yavuyemo nayo izagabanuka.Turasaba cyane ko urugero rwambere rwicyitegererezo rutagomba kurenza mg 50 kubikorwa byo gukuramo RNA.
7.Ijwi ridakwiriye cyangwa gusohora kutuzuye.
Igitekerezo: Ingano yoroheje yinkingi yo kweza ni 50-200 μl;niba ingaruka zo gukuraho zidashimishije, birasabwa kongera igihe mubushyuhe bwicyumba nyuma yo kongeramo ubushyuhe bwa RNase-Free ddH2O, nka 5-10min.
8.Inkingi yo kweza ifite ibisigisigi bya Ethanol nyuma yo gukaraba hamwe na BufferPRW2.
Igitekerezo: Niba umuyoboro wubusa ushyizwemo min min 1 kandi haracyari etanol isigaye nyuma yo koza muri Buffer PRW2, urashobora kongera igihe cya centrifugation yubusa kugeza min 2, cyangwa ugashyira inkingi yo kweza mubushyuhe bwicyumba kuminota 5 kugirango ukureho Ethanol isigaye.
9.Ibikoresho byakoreshejwe nabi.
Icyifuzo: Kubihingwa by'icyitegererezo cya polifenolike polysaccharide, ukoresheje ibikoresho bisanzwe nka Plant Total RNA Isolation Kit ntibishobora kubona urugero rwiza rwa RNA.Turagusaba gukoresha Ikimera Cyuzuye RNA IsolationKit Plus, yagenewe byumwihariko kuburugero rwibiti bya polifenolike polysaccharide.Igikoresho cyakozwe muburyo bwo gukuramo RNA muri polifenol na polysaccharide yintangarugero.
OD260 / OD280 agaciro kari hasi
RNA ikuraho hamwe na ddH2O kandi ikoreshwa mugusoma spekitifotometero ibisubizo mubisubizo bya OD260 / OD280.Turasaba ko dukoresha mM 10 Tris-HCl, pH 7.5 (aho kuba RNase-Free ddH2O kugirango duhindure RNA) kugirango tubone indangagaciro za OD260 / OD280, reba "RNA Kwibanda no Kweza" kurupapuro rwa 19.
RNA yatunganijwe yangiritse
Ubwiza bwa RNA isukuye bujyanye nibintu nko kubungabunga icyitegererezo, kwanduza RNase, no gukoreshwa.
Isesengura ry'impamvu zisanzwe:
1.Icyitegererezo nticyabitswe mugihe cyo gukusanya.
Icyifuzo: Niba ingero za tissue zidakoreshejwe mugihe cyo gukusanya, nyamuneka uzibike muri azote yuzuye mumazi yubushyuhe buke ako kanya cyangwa uyohereze kuri -80 ° C kugirango ubike igihe kirekire nyuma yo gukonjeshwa vuba muri azote yuzuye, cyangwa uhite wibiza ibyitegererezo muri RNA stabilizer RNAlater igisubizo (icyitegererezo cyinyamaswa).Kubikuramo RNA, gerageza gukoresha ingero za tissue nshya zegeranijwe.
2.Gusubiramo kenshi no gukonjesha ingero za tissue.
Icyifuzo: Iyo ubitse ingero za tissue, nibyiza kubicamo uduce duto kugirango tubungabunge, hanyuma ugakuramo igice cyabyo mugihe ubikoresheje kugirango wirinde kwangirika kwa RNA guterwa no gukonjesha no gukonjesha ingero.
3.RNase yatangijwe mucyumba cyo gukoreramo cyangwa ntabwo yambarwa na gants imwe, masike, nibindi.
Icyifuzo: Ubushakashatsi bwo gukuramo RNA bukorwa neza mubikorwa bitandukanye bya RNA, kandi ameza ya laboratoire agomba gusukurwa mbere yubushakashatsi, kandi uturindantoki twajugunywe hamwe na masike bigomba kwambarwa mugihe cyibigeragezo kugirango twirinde kwangirika kwa RNA guterwa no kwinjiza RNase ku rugero runini.
4.Ragent yanduye na RNase mugihe cyo kuyikoresha.
Icyifuzo: Simbuza urukurikirane rushya rwibimera byose RNA ikuramo ibikoresho kubushakashatsi bujyanye.
5.Icyuma cya centrifuge hamwe na pipette inama zikoreshwa muri RNA manipulation zanduye na RNase.
Igitekerezo: Menya neza ko imiyoboro ya centrifuge, inama za pipeti, imiyoboro, nibindi bikoreshwa mugukuramo RNA byose ni RNase-Ubuntu.
Imfashanyigisho:
Gutera Igiteranyo cya RNA Kwigunga Kit Wongeyeho Igitabo
