Jagora don nazarin matsalolin

The following is an analysis of the problems that might be encountered in the extraction of viral RNA. We wish it would be helpful to your experiment. In addition, for other experimental or technical problems other than operating instructions and problem analysis, we have dedicated technical support to help you. Contact us if you need at : 028-83361257or E-mail：Tech@foregene.com。

Ba za a iya fitar da RNA ba ko yawan amfanin acid nucleic ya yi ƙasa

Yawancin lokaci akwai dalilai da yawa waɗanda ke shafar ingancin farfadowa, kamar: samfurin abun ciki na RNA, hanyar aiki, ƙarar haɓakawa, da sauransu.

Binciken dalilai na gama gari:

1.Ice wanka ko ƙananan zafin jiki (4 ° C) centrifugation yayin aiki.

Shawarwari: Yanayin zafin jiki (15-25 ° C) aiki, ba wankan kankara da ƙananan zafin jiki ba.

2. Samfurin da ba daidai ba ko ajiyar samfurin na dogon lokaci.

Shawara: Ajiye samfurori a -80 ° C ko daskare a cikin ruwa nitrogen, kuma kauce wa daskare-narke maimaita amfani;gwada amfani da sabbin samfuran da aka tattara don hakar RNA.

3. Rashin isasshen samfurin lysis

Shawarwari: Da fatan za a tabbatar da cewa samfurin da maganin aiki (Linear Acrylamide) an haɗa su sosai kuma an sanya su na minti 10 a zazzabi na ɗaki (15-25 ° C)

4.An ƙara eluent ba daidai ba

Shawarwari: Tabbatar cewa an ƙara ddH2O-Free RNase zuwa tsakiyar membrane na ginshiƙin tsarkakewa.

5.Ba daidai ba girma na anhydrous ethanol a cikin Buffer viRW2

Shawara: Da fatan za a bi umarnin, ƙara madaidaicin ƙarar ethanol mai anhydrous zuwa Buffer viRW2 kuma a haɗa su da kyau kafin amfani da kit ɗin.

6.Yin amfani da samfur mara kyau.

Shawara: 200µl na samfurin a kowace 500μl na Buffer viRL.Ƙarfin samfurin da ya wuce kima zai haifar da raguwar adadin hakar RNA.

7.Ba daidai ba ƙarar ƙira ko rashin cikawa.

Shawarwari: Ƙaƙwalwar haɓakar ginshiƙin tsarkakewa shine 30-50μl;idan tasirin elution bai gamsar ba, ana ba da shawarar ƙara ddH mai zafi-Free RNase.₂O kuma ƙara lokacin sanyawa a zafin jiki, kamar minti 5-10

8.Purification shafi yana da ragowar ethanol bayan kurkura a cikin Buffer viRW2.

Shawara: Idan har yanzu ethanol ya kasance bayan kurkura a cikin Buffer viRW2 da fanko-tube centrifugation na 2min, za'a iya barin ginshiƙin tsarkakewa a cikin zafin jiki na 5min bayan centrifugation na bututu don cikakken cire sauran ethanol.

Lalacewar ƙwayoyin RNA masu tsafta

Ingancin RNA da aka tsarkake yana da alaƙa da abubuwa kamar ajiyar samfur, gurɓataccen RNase, da aiki.

Binciken dalilai na gama gari:

1.Ba a adana samfurori da aka tattara a lokaci ba.

Shawara: Idan ba a yi amfani da samfurin a cikin lokaci bayan tattarawa ba, da fatan za a adana shi a -80 ℃ ko nitrogen ruwa nan da nan.Don hakar kwayoyin RNA, gwada amfani da samfurori da aka tattara a duk lokacin da zai yiwu.

2. Samfuran da aka tattara suna daskarewa kuma suna narke akai-akai.

Shawara: A guji maimaita daskarewa da narke (ba fiye da sau ɗaya ba) yayin tattara samfuri da adanawa, in ba haka ba yawan adadin acid nucleic zai ragu.

An gabatar da 3.RNase a cikin dakin aiki ko kuma ba a sa safar hannu, abin rufe fuska, da sauransu.

Shawara: An fi yin hakar gwajin kwayoyin RNA a cikin wani dakin aikin RNA daban, kuma ana tsaftace teburin gwajin kafin gwajin.Saka safofin hannu da abin rufe fuska yayin gwajin don guje wa lalatawar RNA ta hanyar gabatarwar RNase.

4.The reagent yana gurbata ta RNase yayin amfani.

Shawara: Sauya da sabon Katin Keɓewar RNA na ƙwayar cuta don gwaje-gwaje masu alaƙa.

5.Cibiyar RNase na bututun centrifuge, tukwici na pipette, da sauransu. Shawara: Tabbatar cewa bututun centrifuge, tukwici na pipette, da pipettes duk ba su da RNase.

Kwayoyin RNA da aka tsarkake sun shafi gwaje-gwaje na ƙasa

Kwayoyin RNA da aka tsarkake ta hanyar ginshiƙin tsarkakewa za su yi tasiri ga gwaje-gwajen da ke ƙasa idan akwai ions gishiri ko sunadaran da yawa, kamar: fassarar juzu'i, Northern Blot, da sauransu.

1.Akwai sauran ions gishiri a cikin kwayoyin RNA da aka eluted.

Shawarwari: Tabbatar cewa an ƙara madaidaicin ƙarar ethanol anhydrous zuwa Buffer viRW2, kuma ku wanke ginshiƙin tsarkakewa sau biyu bisa ga madaidaicin saurin centrifugation akan umarnin aiki;Idan har yanzu akwai sauran ions gishiri, zaku iya ƙara Buffer viRW2 zuwa ginshiƙin tsarkakewa, kuma ku bar shi a cikin dakin zafin jiki na 5min.Sa'an nan kuma yi centrifugation don cire gurɓatar ions gishiri zuwa mafi girma

2.Akwai sauran ethanol a cikin kwayoyin RNA da aka eluted

Shawara: da zarar an tabbatar da cewa Buffer viRW2 ya wanke ginshiƙan tsarkakewa, aiwatar da centrifugation na fanko bisa ga saurin centrifugal akan umarnin aiki.Idan har yanzu akwai sauran ethanol, ana iya barin shi na mintuna 5 a zafin jiki na ɗaki bayan centrifugation na bututu don cire sauran ethanol zuwa mafi girma.

Jagoran Jagora:

Littafin Umarnin Keɓewa na RNA Viral