Siphishekela imfundiso yokuphatha ethi "Ikhwalithi iyingqayizivele, Usizo luphakeme kakhulu, Idumela kuqala", futhi sizodala futhi sabelane ngempumelelo nawo wonke amakhasimende e-China wholesale yase-China Magpure Ffpe Rna KitI-Nucleic Acid Isolation& Ikhithi Yokuhlanza, Sithembisa ukuzama okusemandleni ethu ukukunikeza ngekhwalithi ephezulu nemikhiqizo namasevisi asebenzayo.
Siphishekela imfundiso yokuphatha ethi “Ikhwalithi iyingqayizivele, Usizo luphakeme, Idumela kuqala”, futhi sizodala futhi sabelane ngempumelelo nawo wonke amakhasimendeChina Rna/DNA Isizinda Ukuhlanzwa, I-Nucleic Acid Isolation, Ngohlelo lokusebenza oludidiyelwe ngokugcwele, inkampani yethu izuze udumo oluhle ngempahla yethu esezingeni eliphezulu, amanani aphusile nezinkonzo ezinhle.Ngaleso sikhathi, manje sesisungule uhlelo oluqinile lokuphatha ikhwalithi olwenziwa ezintweni ezingenayo, ezicutshungulwayo kanye nokulethwa.Ngokuhambisana nomgomo othi “Ikhredithi kuqala kanye nokuphakama kwekhasimende”, samukela ngobuqotho amakhasimende avela ekhaya naphesheya ukuze abambisane nathi futhi athuthuke ndawonye ukuze sakhe ikusasa eliqhakazile.

Imininingwane

50 Preps, 200 Preps Ikhithi isebenzisa ikholomu yokuphotha kanye nefomula eyakhiwe yi-Foregene, engakhipha ngokuphumelelayo inani eliphelele le-RNA esezingeni eliphakeme nekhwalithi ephezulu kumathishu ezitshalo ahlukahlukene ane-polysaccharides ephezulu noma okuqukethwe kwama-polyphenols.Ihlinzeka ngekholomu ye-DNA-Cleaning engasusa kalula i-genomic DNA ku-supernatant kanye ne-tissue lysate.Ikholomu ye-RNA kuphela ingabopha ngempumelelo i-RNA.Ikhithi ingacubungula inani elikhulu lamasampuli ngesikhathi esisodwa.

Lonke uhlelo aluqukethe i-RNase, ngakho-ke i-RNA ehlanziwe ngeke yehliswe.I-Buffer PRW1 kanye ne-Buffer PRW2 ingaqinisekisa ukuthi i-RNA etholiwe ayingcoliswanga amaprotheni, i-DNA, ama-ion, nezinhlanganisela eziphilayo.

Izingxenye zekhithi

Izici&izinzuzo

■ Ukusebenza ekamelweni lokushisa (15-25℃) kuyo yonke inqubo, ngaphandle kokugeza eqhweni nokushisa okuphansi kwe-centrifugation.■ Ikhithi ephelele ye-RNase-Free, asikho isidingo sokukhathazeka ngokuwohloka kwe-RNA.■ Ifaneleka ngokukhethekile ukuhlanzwa kwe-RNA kumasampula ezitshalo ama-polysaccharides nama-polyphenols.■ Ikholomu Yokuhlanza I-DNA ibophezela ngokukhethekile ku-DNA, ukuze ikhithi isuse ukungcoliswa kwe-DNA ye-genomic ngaphandle kokwengeza i-DNase.■ Isivuno esikhulu se-RNA: Ikholomu ye-RNA kuphela kanye nefomula eyingqayizivele ingahlanza i-RNA ngokuphumelelayo.■ Isivinini esisheshayo: kulula ukusebenza futhi singaqedwa phakathi nemizuzu engama-30.■ Ukuphepha: asikho i-reagent ephilayo edingekayo.■ Ikhwalithi ephezulu: Izingcezwana ze-RNA ezihlanziwe zimsulwa kakhulu, azinawo amaphrotheni nokunye ukungcola, futhi zingahlangabezana nezinhlelo zokuhlola ezihlukahlukene ezansi nomfula.

Imingcele yomkhiqizo

■ Izicelo eziwela phansi: ukuhlanganiswa kwe-cDNA yomucu wokuqala, i-RT-PCR, i-molecular cloning, i-Northern Blot, njll. ■ Isampula: Izicubu zezitshalo ezintsha noma eziqandisiwe zama-polysaccharides nama-polyphenols ■ Umthamo: 50mg izicubu zesitshalo ■ Umthamo omkhulu wokubopha i-RNA wekholomu yokuhlanza: 80 μg 20 ■ 50 Elution

Isicelo sekhithi

Ilungele ukukhishwa kanye nokuhlanzwa kwe-RNA ephelele kumasampula ezicubu zezitshalo ezintsha noma eziqandisiwe (ikakhulukazi izicubu zamaqabunga ezitshalo ezintsha) ezinokuqukethwe okuphezulu kwe-polysaccharide kanye ne-polyphenol.

Ukugeleza komsebenzi

Umdwebo

I-Plant Total RNA Isolation Kit Plus icutshungulwe u-50mg wamaqabunga amasha e-polysaccharides nama-polyphenols, futhi i-RNA ehlanzekile engu-5% yahlolwa nge-electrophoresis.1: Ubhanana 2: I-Ginkgo 3: Ukotini 4: Ihalananda

Isitoreji kanye Nempilo yeshelufu

Le kit ingagcinwa izinyanga ezingu-24 ngaphansi kwezimo ezomile ekamelweni lokushisa (15-25℃);uma idinga ukugcinwa isikhathi eside, ingagcinwa ku-2–8℃.I-Buffer PSL1 ingafakwa ku-4℃ inyanga engu-1 ngemva kokwengeza i-β-mercaptoethanol (kuyanconywa ukuthi uyengeze ngesikhathi esifanayo sokuhlola). Siphishekela imfundiso yokuphatha yokuthi "Ikhwalithi iyingqayizivele, Usizo luphakeme kakhulu, Isithunzi siqala", futhi sizodala futhi sabelane ngempumelelo nawo wonke amaklayenti e-China wholesalef China MagpuretI-Nucleic Acid Isolation& Ikhithi Yokuhlanza, Sithembisa ukuzama okusemandleni ethu ukukunikeza ngekhwalithi ephezulu nemikhiqizo namasevisi asebenzayo.
China wholesaleChina Rna/DNA Isizinda Ukuhlanzwa, I-Nucleic Acid Isolation, Ngohlelo lokusebenza oludidiyelwe ngokugcwele, inkampani yethu izuze udumo oluhle ngempahla yethu esezingeni eliphezulu, amanani aphusile nezinkonzo ezinhle.Ngaleso sikhathi, manje sesisungule uhlelo oluqinile lokuphatha ikhwalithi olwenziwa ezintweni ezingenayo, ezicutshungulwayo kanye nokulethwa.Ngokuhambisana nomgomo othi “Ikhredithi kuqala kanye nokuphakama kwekhasimende”, samukela ngobuqotho amakhasimende avela ekhaya naphesheya ukuze abambisane nathi futhi athuthuke ndawonye ukuze sakhe ikusasa eliqhakazile.

Ikholomu ixhunyiwe

Ngemuva kokuthi ikholomu ixhunyiwe, isivuno se-RNA siyancishiswa noma akunakwenzeka ukuhlanza i-RNA, futhi isisindo esitholiwe se-RNA siphansi.

Ukuhlaziywa kwembangela evamile:

1. Ukuphumula kwesampula akuphelele.

Ukuphuka kwesampula akubangeli ngokuphelele i-DNA-CLEANING COLUMN ukuthi ivinjwe, kuyilapho kuthinta isivuno nekhwalithi ye-RNA.Sincoma ukusebenza kokugaya ngokushesha ku-nitrogen ewuketshezi eyanele lapho uphula amasampula, Zama ukuchoboza udonga lwamaseli wesampula, ulwelwesi lwamaseli nezinye izicubu.Kumasampula ezitshalo e-polysaccharides, sincoma ukuthi usebenzise I-Plant Total RNA IsolATION KIT PLUS.

2. Uma umunca isampula ye-supernatant ehlukanisiwe Ngekholomu Yokuhlanza Ye-DNA, i-precipitate yeseli engaba yizicucu ingase ihogelwe.

Izinsalela ezihlukene zeseli ezithathiwe zizobangela Ikholomu YE-RNA KUPHELA ezovinjwa lapho kwenziwa umsebenzi wokukhangisa we-RNA (bona isinyathelo 6).Sincoma ukuthi uqaphele lapho uncela lesi sidakamizwa esinamandla ukuze ugweme ukumuncwa ama-cell debris.

3. Inani lokuqala lesampula likhulu kakhulu.

Ukusetshenziswa kwesampula okweqile kuzoholela ekuhlukaneni kwesampula okungaphelele noma ukuguqulwa kweseli okungaphelele nge-Buffer PSL1, okuholela ekuvinjweni kwekholomu yokuhlanza ngesikhathi sokuhlanza.Ikhithi Yokuhlukanisa Yezitshalo Isamba se-RNA Isampula ngayinye yokusebenza ehlanziwe ingu-50 mg.Ngamasampula ezitshalo e-polysaccharides, sincoma ukuthi uzame I-Plant Total RNA IsolATION KIT PLUS.

4. Izinga lokushisa le-centrifuge liphansi kakhulu.

Yonke inqubo yokuhlukaniswa kwe-RNA kanye nenqubo yokuhlanza yenziwa ekamelweni lokushisa (20-25°C), ngaphandle kokuthi isicubu sesampula siphulwa yi-nitrogen ewuketshezi.Izinga lokushisa kwamanye ama-cryogenic centrifuges lingaphansi kuka-20℃, okungase kubangele ukuvinjwa Kwekholomu Yokuhlanza I-DNA kanye/noma Ikholomu Ye-RNA Kuphela.Uma lokhu kwenzeka, setha izinga lokushisa le-centrifuge libe ngu-20-25℃, futhiqiniseka ukuthi ingxube ye-lysis kanye/noma i-ethanol-added supernatant ishiselwe kuze kube ngu-37°C.

Ayikho i-RNA ekhishiwe noma isivuno se-RNA siphansi

Ngokuvamile kuba nezinto eziningi ezithinta ukusebenza kahle kokuthola kabusha, njengalezi: isampula yokuqukethwe kwe-RNA, indlela yokusebenza, ivolumu ye-elution, njll.

Ukuhlaziywa kwezimbangela ezivamile njengoba ngezansi:

I-1.Ukugeza kweqhwa noma izinga lokushisa eliphansi (4 ° C) i-centrifugation yenziwa ngesikhathi sokusebenza.

Ukusikisela: Isebenza ekamelweni lokushisa (15-25°C) kuyo yonke inqubo, ungenzi ukugeza kweqhwa kanye ne-centrifugation yokushisa ephansi.

2.I-RNA yehlisiwe ngenxa yokulondolozwa okungafanele kwesampula noma ukulondolozwa kwesikhathi eside kwesampula.

Okutuswayo: Amasampula asanda kuqoqwa kufanele aqandwe ngokushesha ku-nitrogen ewuketshezi, abese egcinwa ku -80°C isikhathi eside, agweme ukuqhwaza okuphindaphindiwe nokuncibilika kwamasampuli;noma ngokushesha cwilisa amasampula ku-RNA stabilizer RNAlater solution (amasampula ezilwane).

3.Ukuhlukaniswa kwesampula okunganele kanye ne-lysis kuholela ekuvinjweni kwekholomu yokuhlanza.

Isiphakamiso: Uma ugaya izicubu, sicela uqinisekise ukuthi isicubu sigaywe ngokwanele, futhi usidlulisele ngokushesha ku-Buffer PSL1 esilungiselelwe ngaphambilini (qinisekisa ukuthi ingxenye elungile ye-β-ME yengeziwe, bona isinyathelo 1 senqubo).

4.I-eluent yengezwe ngokungalungile.

Isiphakamiso: Qiniseka ukuthi i-ddH2O ye-RNase-Free iconsiselwa phakathi nolwelwesi lwekholomu yokuhlanza.

5.Ivolumu elungile ye-ethanol ephelele ayizange yengezwe ku-Buffer PSL2 noma ku-Buffer PRW2.

Ukusikisela: Sicela ulandele imiyalelo, wengeze ivolumu efanele ye-ethanol ephelele ku-Buffer PSL2 kanye ne-Buffer PRW2 bese uxuba kahle ngaphambi kokuba ikhithi isetshenziswe.

6.Inani lesampula lethishu alifanelekile.

Isiphakamiso: Sebenzisa u-50 mg wethishu ngo-500 μl we-Buffer PSL1.Ukusebenzisa izicubu eziningi kuzonciphisa inani le-RNA ekhishiwe futhi ukuhlanzeka kwe-RNA okuwumphumela nakho kuzoncipha.Sincoma ngokuqinile ukuthi umthamo wokuqala wesampula akufanele udlule u-50 mg ngokusebenza kokukhipha kwe-RNA ngakunye.

7.Ivolumu ye-elution engafanele noma ukungezwani okuphelele.

Isiphakamiso: Umthamo ongacacile wekholomu yokuhlanza ungama-50-200 μl;uma umphumela we-elution ungagculisi, kuyanconywa ukuthi unwebe isikhathi kuzinga lokushisa legumbi ngemva kokwengeza i-ddH2O eshisiwe ye-RNase-Free, njenge-5-10min.

8.Ikholomu yokuhlanza inensalela ye-ethanol ngemva kokugeza nge-BufferPRW2.

Isiphakamiso: Uma ishubhu elingenalutho li-centrifuged iminithi elingu-1 futhi kusekhona i-ethanol esele ngemva kokugeza ku-Buffer PRW2, ungakwazi ukwandisa isikhathi se-tube centrifugation engenalutho sibe yimizuzu emi-2, noma ubeke ikholomu yokuhlanza ekamelweni lokushisa imizuzu emi-5 ukuze ususe ngokuphelele i-ethanol esele.

9.Ikhithi isetshenziswe ngokungalungile.

Isiphakamiso: Kumasampula ezitshalo we-polyphenolic polysaccharides, ukusebenzisa amakhithi avamile njenge-Plant Total RNA Isolation Kit kungase kungakwazi ukuthola amasampula e-RNA afanelekile.Sincoma ukuthi usebenzise i-Plant Total RNA IsolationKit Plus, eklanyelwe ngokukhethekile amasampula ezitshalo ze-polyphenolic polysaccharide.Ikhithi eyakhelwe ngokukhethekile ukukhipha i-RNA ku-polyphenol namasampula ezitshalo ze-polysaccharide.

OD260/OD280 inani liphansi

I-RNA elution ene-ddH2O futhi isetshenziselwa ukufundwa kwe-spectrophotometer iphumela kumanani aphansi e-OD260/OD280.Sincoma ukusebenzisa u-10 mM Tris-HCl, pH 7.5 (kunokuba i-RNase-Free ddH2O ukuze ukhiphe i-RNA) ukuze uthole amanani alungile uma kuqhathaniswa e-OD260/OD280, bona “I-RNA Concentration and Purification Assays” ekhasini 19.

I-RNA ehlanziwe yehlisiwe

Ikhwalithi ye-RNA ehlanzekile ihlobene nezici ezifana nokulondolozwa kwesampula, ukungcoliswa kwe-RNase, nokukhohlisa.

Ukuhlaziywa kwezimbangela ezijwayelekile:

1.Amasampula ezicubu awazange agcinwe ngesikhathi ngemuva kokuqoqwa.

Okutuswayo: Uma amasampula ezicubu engasetshenziswa ngesikhathi ngemva kokuqoqwa, sicela uwagcine ku-nitrogen ewuketshezi ezingeni lokushisa eliphansi ngokushesha noma uwadlulisele ku--80°C ukuze agcinwe isikhathi eside ngemva kokuqhwaza okusheshayo ku-nitrogen ewuketshezi, noma ngokushesha ucwilise amasampula ku-RNA stabilizer RNAlater solution (amasampula ezilwane ).Ukuze uthole i-RNA, zama ukusebenzisa amasampula ezicubu ezisanda kuqoqwa.

2.Ukuqhwaza okuphindaphindiwe nokuncibilika kwamasampula ezicubu.

Ukusikisela: Lapho ugcina amasampula ezicubu, kungcono ukuwasika abe izingcezu ezincane ukuze alondolozwe, futhi ukhiphe ingxenye yawo lapho uwasebenzisa ukuze ugweme ukuwohloka kwe-RNA okubangelwa ukubanda okuphindaphindiwe nokuncibilika kwamasampula.

3.I-RNase yethulwa egunjini lokuhlinza noma engagqoki amagilavu ​​alahlwayo, imaski, njll.

Isiphakamiso: Ukuhlolwa kokukhipha i-RNA kwenziwa kangcono kakhulu emisebenzini ehlukene ye-RNA, futhi ithebula laselabhorethri kufanele lihlanzwe ngaphambi kokuhlolwa, futhi amagilavu ​​alahlwayo namamaski kufanele kugqokwe phakathi nokuhlolwa ukuze kugwenywe ukuwohloka kwe-RNA okubangelwa ukwethulwa kwe-RNase ngezinga elikhulu kakhulu.

4.I-reagent ingcoliswe yi-RNase ngesikhathi sokusetshenziswa.

Isiphakamiso: Faka esikhundleni ngochungechunge olusha lwamakhithi okukhipha i-RNA esitshalo esiphelele sokuhlolwa okuhlobene.

5.Amashubhu e-centrifuge namathiphu we-pipette asetshenziselwa ukukhwabanisa kwe-RNA angcoliswe yi-RNase.

Ukusikisela: Qinisekisa ukuthi amashubhu e-centrifuge, amathiphu e-pipette, ama-pipette, njll. asetshenziswa ekukhishweni kwe-RNA konke Akuna-RNAse.