Uhlobo: Spin ikholomu isuselwe
Isampula: I-Plasma, i-serum, uketshezi lomzimba olungenaselula, njll.
Ithagethi: Igciwane le-RNA.
Iqala ivolumu: ~ 140 μl
Isikhathi sokusebenza: ~ 1 ihora
Izinhlelo zokusebenza ezisezansi: I-RT-PCR / RT-qPCR, ukwakhiwa kwelabhulali ye-NGS, ukusabela kwe-enzyme, njll.
A-1 Ukuhlushwa okuphansi kwamaseli noma igciwane kusampula yokuqala
-Nothisa ukucinana kwamaseli noma kwamagciwane.
I-A-2 I-lysis enganele yamasampuli
—Amasampula awakaxutshwa kahle ne-lysis buffer. Kuphakanyiswa ukuthi uhlanganise kahle nge-pulse-vortexing izikhathi eziyi-1-2.
—I-lysis enganele yeseli ebangelwa ukwehla komsebenzi we-proteinase K.
—Ukulimala kwamaseli noma ukonakala kwamaprotheni ngenxa yesikhathi esanele sokugeza. Kuphakanyiswa ukusika izicubu zibe yizicucu ezincane bese unweba isikhathi sokugeza ukuze ususe yonke insalela ku-lysate.
I-A-3 Ukukhangisa okwanele kwe-DNA.
-Ayikho i-ethanol noma amaphesenti aphansi esikhundleni se-100% ethanol engezwe ngaphambi kokuthi i-lysate idluliselwe kukholamu ye-spin.
A-4 Inani le-pH le-elution buffer liphansi kakhulu.
- Lungisa i-pH iye phakathi kuka-8.0-8.3.
I-ethanol eyinsalela ku-eluent.
—Kunensalela yokuwasha i-PW esikhaleni. I-ethanol ingasuswa ngokufaka phakathi ikholamu ye-spin ngemizuzu engu-3-5, bese ifakwa ekamelweni lokushisa noma ku-50 ℃ incubator ye-1-2 min.
A-1 Isampula akuyona entsha.
—Khipha i-DNA eyisampula enhle njengokulawula ukunquma ukuthi ngabe i-DNA esampulweni yehle yini.
Ukwelashwa kwangaphambili okungalungile.
—Kudalwa ukugaya okweqile kwe-nitrogen, uketshezi olubuyisa umswakama, noma inani elikhulu kakhulu lesampula.
Ama-pretreatment kufanele ahluke kumasampula ahlukile. Ngamasampula ezitshalo, qiniseka ukuthi ugaya kahle i-nitrogen engamanzi. Ngamasampuli ezilwane, homogenate noma gaya kahle i-nitrogen engamanzi. Kumasampuli anezindonga zamaseli okunzima ukugqashuka, njengama-bacterium e-G + nemvubelo, kuphakanyiswa ukusebenzisa i-lysozyme, i-lyticase noma izindlela zemishini yokwephula izindonga zeseli.
I-4992201/4992202 iPlant Genomic DNA Kit isebenzisa indlela esekwe kukholamu edinga i-chloroform yokukhipha. Ilungele ikakhulukazi amasampula ezitshalo ezahlukahlukene, kanye ne-powder eyomile yesitshalo. IHi-DNAsecure Plant Kit nayo isekelwe kwikholomu, kepha asikho isidingo sokukhishwa kwe-phenol / chloroform, okuyenza iphephe futhi ingabi nobuthi. Ifanele izitshalo ezinama-polysaccharides aphezulu nokuqukethwe kwe-polyphenol. I-4992709/4992710 DNAquick Plant System isebenzisa indlela esekelwa uketshezi. Ukukhishwa kwePhenol / chloroform akudingeki futhi. Inqubo yokuhlanzwa ilula futhi iyashesha ngaphandle komkhawulo wamanani okuqala esampula, ngakho-ke abasebenzisi bangalungisa inani ngokuvumelana nezidingo zokuhlola. Usayizi omkhulu wezingcezu ze-gDNA ungatholakala ngesivuno esikhulu.
I-genomic DNA ikhishwe emananini ahlukene amasampuli egazi lomuntu yi-TIANamp Blood DNA Kit. Imiphumela imi kanje. Imiphumela ibalwe njengereferensi kuphela, imiphumela yangempela yokukhipha incike ezimweni zamasampuli.
Ukukhishwa kwegazi kwe-DNA kwegazi kungenziwa kusetshenziswa ama-reagents ahlinzekwe kulawa makhithi amabili ngokumane uguqule umthetho olandelwayo uye emyalweni othize wokukhishwa kwegazi kwe-DNA. Ikhophi ethambile ye-clot yegazi ye-DNA protocol ingakhishwa uma ucela.
Misa okwesikhashana isampula elisha nge-1 ml PBS, usawoti ojwayelekile noma i-TE buffer. Homogenize Ngokuphelele isampula nge-homogenizer bese uqoqa imvula phansi kwephubhu nge-centrifuging. Lahla amandla angaphezu kwawemvelo, uphinde uvuselele ukugqashuka nge-200 μl buffer GA. Ukuhlanzwa kwe-DNA okulandelayo kungenziwa ngokuya ngemiyalo.
Ukuhlanzwa kwe-gDNA kumasampula e-plasma, serum kanye ne-fluid fluid, Kunconywa i-TIANamp Micro DNA Kit. Ukuhlanzwa kwegciwane i-gDNA kusuka kumasampula we-serum / plasma, Kunconywa i-TIANamp Virus DNA / RNA Kit. Ukuhlanzwa kwe-gDNA yebhaktheriya kusuka kumasampula we-serum ne-plasma, i-TIANamp Bacteria DNA Kit inconywa (i-lysozyme kufanele ifakwe ibhaktheriya elihle). Kumasampula amathe, i-Hi-Swab DNA Kit ne-TIANamp Bacteria DNA Kit kuyanconywa.
I-DNAsecure Plant Kit noma i-DNAquick Plant System inconyelwa ukukhishwa kwefungal genome. Ngokukhishwa kwemvubelo ye-genome, Kunconywa i-TIANamp Yeast DNA Kit (i-lyticase kufanele izilungiselele).
Yonke imikhiqizo ingenziwa ngezifiso i-ODM / OEM. Ngemininingwane,sicela uchofoze i-Customized Service (ODM / OEM)
I-AIV-H5 yahlanjululwa yaba yi-10-5-10-8ama-gradients ngamanzi we-Mill-Q. Isampula le-140 μl likhishwe yi-TIANamp Virus RNA Kit (DP315-R) kanye nekhithi efanelekile evela kumhlinzeki A. Kuqhathaniswa noMhlinzeki A, I-TIANGEN inokusebenza okuphezulu kwesampula lokuhlushwa eliphansi.
Ikhithi yokuthola i-RT-qPCR: TIANGEN, FastKing One Step RT-qPCR kit (Probe)
Insimbi ye-RT-qPCR: Uhlelo lwe-PCR olusheshayo lwangempela lwe-AB 7500.
Kusukela yasungulwa, imboni yethu ibilokhu ithuthukisa imikhiqizo esezingeni lomhlaba yokuqala ngokunamathela esimisweni
wekhwalithi kuqala. Imikhiqizo yethu izuze idumela elihle kakhulu embonini futhi ithembekile phakathi kwamakhasimende amasha namadala ..