Jimlar Dabbobin Keɓewar RNA Jimlar Cirar RNA da Kayan Tsabtace don Naman Dabbobi & Kwayoyin Halitta
Bayanin Kit:
Cikin sauri da inganci cire tsafta mai inganci da jimillar RNA daga kyallen dabbobi daban-daban.
Babu buƙatar damuwa game da lalata RNA.Duk tsarin ba shi da RNase-Free
Cire DNA yadda yakamata ta amfani da Rukunin Tsabtace DNA
Cire DNA ba tare da ƙara DNA ba
Sauƙaƙe-dukkan ayyukan ana kammala su a cikin zafin jiki
Ana iya kammala aiki mai sauri a cikin mintuna 30
Amintacciya — ba a yi amfani da reagent Organic ba
Babban tsarki-OD260/280≈1.8-2.1
Bayanin Kits
Shirye-shirye 50, Shirye-shiryen 200
Wannan kit yana amfani da shijuya shafi da dabarawanda kamfaninmu ya haɓaka, wanda zai iya fitar da RNA mai tsabta da inganci mai kyau daga nau'ikan nau'ikan dabbobi daban-daban tare da ingantaccen aiki.Yana samar da ingantaccen ginshiƙi na DNA-Cleaning, wanda zai iya raba sauƙi da kuma tallata DNA na genomic daga supernatant da lysate nama, mai sauƙi da ceton lokaci;Rukunin RNA-kawai na iya ɗaure RNA da kyau da kyau kuma ana iya sarrafa su lokaci guda tare da keɓancewar dabara Yawancin samfurori.
Dukkanin tsarin ba shi da RNase-Free, don kada RNA da aka fitar ba ta lalacewa ba;Buffer RW1, Tsarin wanke buffer RW2, ta yadda RNA da aka samu ba ta da furotin, DNA, ion, da gurɓataccen mahalli.
Abubuwan Kit
| Jimlar Dabbobin Keɓewar RNA | ||
| Abubuwan Kit | RE-03011 | RE-03014 |
| 50 T | 200 T | |
| RL1* | ml 25 | 100 ml |
| Farashin RL2 | ml 15 | ml 60 |
| RW1* | ml 25 | 100 ml |
| Farashin RW2 | 24ml ku | ml 96 |
| RNase-Free ddH2O | ml 10 | ml 40 |
| Rukunin RNA-kawai | 50 | 200 |
| Rukunin Tsabtace DNA | 50 | 200 |
| Jagoran Jagora | guda 1 | guda 1 |
Bayanin samfur
| Tsarin | Juya shafi | Bangaren tsarkakewa | Rukunin Foregene, reagent |
| Flux | 1-24 samfurori | Lokaci kowane shiri | ~ 30 min (samfurori 24) |
| Centrifuge | Tebur centrifuge | Pyrolysis rabuwa | Rabuwar Centrifugal |
| Misali | Naman dabba;tantanin halitta | Adadin samfuran | Nama: 10-20 MG;Cell: (1-5)×106 |
| Ƙarar hasashe | 50-200 ml | Matsakaicin ƙarar lodi | 850 ml |
Fasaloli & fa'idodi
■ Babu buƙatar damuwa game da lalacewar RNA;Duk tsarin ba shi da RNase-Free
∎ Cire DNA da kyau ta amfani da Rukunin Tsabtace DNA
■ Cire DNA ba tare da ƙara DNA ba
■ Ana kammala ayyuka masu sauƙi-duk a yanayin zafin ɗaki
■ Ana iya kammala aikin gaggawa cikin mintuna 30
∎ Amintacciya-babu reagent na halitta da ake buƙata
■ Babban tsarki -OD260/280≈1.8-2.1
Kit aikace-aikace
Ya dace da hakar da tsarkakewar jimillar RNA daga sabo ko daskararru iri-iri na kyallen dabbobi ko sel masu al'ada.
Siffofin samfur
∎ Aikace-aikace na ƙasa: haɗin cDNA na farko, RT-PCR, cloning na kwayoyin halitta, Northern Blot, da sauransu.
Samfurori: kyallen jikin dabba, ƙwayoyin al'ada
■ Adadin: Nama 10-20mg, Kwayoyin (2-5) × 106
■ Ƙarfin ɗaurin DNA na ginshiƙin tsarkakewa: 80 μg
■ Ƙarfin haske: 50-200 μl
zane
Jimlar Kayan Keɓewar Dabbobin RNA da aka yi maganin 20mg
Sabbin samfuran linzamin kwamfuta, ɗauki 5% tsarkakakken jimlar RNA 1% agar
Glycogel electrophoresis
1: Baki 2: Koda
3: Hanta 4: Zuciya
Adana da rayuwar shiryayye
Ana iya adana kit ɗin na tsawon watanni 24 a zazzabi na ɗaki (15-25 ℃) ko 2-8 ℃ na tsawon lokaci.Ana iya adana buffer RL1 a 4 ℃ na wata 1 bayan ƙara β-mercaptoethanol (na zaɓi).
Labarai da aka kawo
1.Idan: 18.808:Zheng, Q., Qin, F., Luo, R., da al.mRNA-Loaded Lipid-Kamar Nanoparticles don Gyara Tushen Hanta Ta Hanyar Haɓaka Ƙirƙirar Ƙirƙirar Ƙirar Tsakiya.Adv.AyyukaMatar.2021, 31, 2011068.doi:10.1002/adfm.202011068.
2.Idan: 18.187:He X, Hong W, Yang J, et al.Kwatsam apoptosis na sel a cikin shirye-shiryen ƙwayar ƙwayar cuta yana yin tasirin immunomodulatory ta hanyar sakin phosphatidylserine.Canjin Siginar Target Ther.2021 Yuli 14; 6 (1): 270.doi: 10.1038/s41392-021-00688-z.
3.Idan: 17.97: Dai Z, Liu H, Liao J, et al.N7-Methylguanosine tRNA gyare-gyare yana haɓaka fassarar mRNA oncogenic kuma yana haɓaka ci gaban cholangiocarcinoma na ciki.Mol Cell.2021 Jul 29: S1097-2765(21)00555-4.doi: 10.1016/j.molcel.2021.07.003.
4.Idan: 9.225: Cao X, Shu Y, Chen Y, et al.Mettl14-Matsakaici m6A Canjin Yana Sauƙaƙe Farfaɗowar Hanta ta hanyar Kula da Endoplasmic Reticulum Homeostasis.Cell Mol Gastroenterol Hepatol.2021;12 (2): 633-651.doi: 10.1016/j.jcmgh.2021.04.001.
RNA ware kayan aikin don sauran samfurin kafofinakwai:
Kwayoyin halitta, shuka, kwayar cuta, jini, da dai sauransu.
Ba a fitar da RNA ko amfanin RNA yayi ƙasa
Sau da yawa akwai dalilai iri-iri waɗanda ke shafar ingancin farfadowa, kamar: samfurin nama na RNA abun ciki, hanyar aiki, ƙarar haɓaka, da sauransu.
1. Ice wanka ko cryogenic (4 ° C) centrifugation aka yi a lokacin aiki.
Shawarwari: Yi aiki a dakin da zafin jiki (15-25 ° C) a duk tsawon tsari, kada ku yi wanka da centrifuge a ƙananan yanayin zafi.
2. Ƙimar samfurin da ba daidai ba ko yawan lokacin ajiyar samfurin.
Shawarwari: Ajiye samfurori a -80 ° C ko daskare a cikin ruwa nitrogen kuma kauce wa daskare-narke maimaita amfani;yi ƙoƙarin amfani da sabobin nama ko ƙwayoyin al'ada don hakar RNA.
3. Rashin isasshen samfurin lysis.
Shawarwari: Lokacin da aka haɗa nama, tabbatar da cewa nama ya isa daidai kuma cewa ƙwayoyin nama sun rabu sosai don bayyana sakin RNA.
4. Ba a ƙara eluent daidai ba.
Shawarwari: Tabbatar da cewa RNase-Free ddH2O ana ƙara juzu'i zuwa tsakiyar membrane ginshiƙin tsarkakewa.
5. Ba a ƙara madaidaicin ƙarar cikakken ethanol zuwa Buffer RL2 ko Buffer RW2 ba.
Shawarwari: Bi umarnin, ƙara madaidaicin ƙarar cikakkar ethanol zuwa Buffer RL2 da Buffer RW2 kuma gauraya da kyau kafin amfani da kit ɗin.
6. Nama samfurin sashi bai dace ba.
Shawarwari: Yi amfani da 10-20 MG na nama ko (1-5) × 106Kwayoyin a cikin 500 μl buffer RL1, saboda yawan amfani da nama zai iya haifar da raguwar hakar RNA.
7. Rashin ingantaccen ƙarar ƙyalli ko rashin cikawa.
Shawarwari: Girman haɓakar ginshiƙin tsarkakewa shine 50-200 μl;idan tasirin elution bai gamsar ba, ana ba da shawarar tsawaita lokacin sanyawa dakin zafin jiki bayan ƙara preheated RNase-Free ddH2O, misali na 5-10 min.
8.Shafin tsarkakewa yana da ragowar ethanol bayan wankewar buffer RW2.
Shawarwari: Idan akwai ragowar ethanol bayan wankewar Buffer RW2, centrifugation mara amfani na 1min, lokacin aikin centrifugation na bututu mara kyau za'a iya ƙara zuwa 2min, ko kuma ana iya sanya ginshiƙin tsarkakewa a dakin da zafin jiki na 5 min don cire isasshen ethanol.
RNA mai tsafta ya lalace
Ingancin RNA da aka tsarkake yana da alaƙa da abubuwa kamar adana samfurin, gurɓataccen RNase, da magudi, da sauransu.
1. Ba a adana samfuran nama a cikin lokaci.
Shawarwari: Idan ba a yi amfani da samfurori na nama ko sel a cikin lokaci ba bayan tattarawa, nan da nan a adana a -80 ° C ko nitrogen mai ruwa.Don cire RNA, yi amfani da sabon samfurin nama ko tantanin halitta a duk lokacin da zai yiwu.
2. Maimaita daskare-narke samfuran nama.
Shawarwari: Lokacin adana samfuran nama, yana da kyau a yanke su kanana don adanawa, kuma a cire ɗaya daga cikin guntun lokacin amfani da su don guje wa daskare-narke samfurin da kuma lalata RNA.
3. Ana gabatar da RNase ko a'a sanye da safar hannu, abin rufe fuska, da sauransu yayin aikin.
Shawarwari: Gwajin cirewar RNA an fi yin su a cikin ɗakunan sarrafa RNA daban kuma an share teburin kafin gwajin.
Sanya safar hannu da abin rufe fuska yayin gwajin don rage lalata RNA wanda ya haifar da gabatarwar RNase.
4. Reagents sun gurbata da RNase yayin amfani.
Shawarwari: Sauya da sabon Kit ɗin Keɓewar Dabbobi na RNA don gwaje-gwaje masu alaƙa.
5. Bututun centrifuge, tukwici, da sauransu da ake amfani da su wajen sarrafa RNA sun gurɓata da RNase.
Shawarwari: Tabbatar da cewa bututun centrifuge, tukwici, pipettes, da sauransu. da ake amfani da su wajen cire RNA duk ba su da RNase.
RNA da aka tsarkake yana shafar gwaje-gwaje na ƙasa
RNA da aka tsarkake ta hanyar ginshiƙin tsarkakewa, idan ions gishiri, abun cikin furotin ya yi girma zai shafi gwajin ƙasa, kamar: juzu'i na baya, Northern Blot et al.
1. RNA da aka elutioned yana da ragowar ion gishiri.
Shawarwari: Tabbatar da cewa an ƙara madaidaicin ƙarar ethanol zuwa Buffer RW2 kuma kuyi ginshiƙan tsarkakewa guda 2 a saurin centrifugal da aka nuna don aiki;idan akwai ragowar ion gishiri, bar ginshiƙin tsarkakewa zuwa Buffer RW2 na 5 min a zafin daki kuma yi centrifugation don haɓaka kawar da gurɓataccen gishiri.
2. Ethanol saura a cikin elutioned RNA.
Shawarwari: Tabbatar da cewa bayan buffer RW2 wankewa, yi aikin centrifugation mara amfani a cikin saurin centrifugation da aka nuna don aiki, ƙara lokacin aikin centrifugation mara amfani zuwa 2 min idan har yanzu akwai ragowar ethanol, ko barin shi a cikin dakin da zafin jiki na 5 min bayan fanko tube centrifugation don ƙara yawan cirewar ethanolue.
