Le tampu isetyenziselwa ukuhlukana kwe-DNA ye-electrophoresis
I-TBE buffer iyincedo ngokukhethekileyo ekuhlukaniseni iincinci ze-DNA ezincinane (MW <1000), ezifana nemveliso emincinci yokukhutshwa kwe- enzyme digest.
I-TBE inomthamo omkhulu wokuxhatshazwa kwaye iya kunika isisombululo esinzulu ngaphezu kwe- TAE buffer . I-TAE buffer isisombululo esenziwe nge-Tris base, acetic acid kunye ne-EDTA (iTris-acetate-EDTA).
I-TBE ibiza ngokugqithiseleyo kune-TAE kwaye inqanda i-DNA ligase, enokubangela iingxaki ukuba kulandelelwe i-DNA yokucocwa nokunyuswa kwamanyathelo. Ngeendlela ezintathu ezilula ezilandelayo, funda indlela yokwenza i-TBE buffer. Akufanele kuthathe ngaphezu kwemizuzu engama-30 ukudala le khowudi. Nantsi indlela:
Lungisa iSolution Solution ye-EDTA
Isisombululo se-EDTA (ethylenediamine tetraacetic acid) kufuneka silungiselelwe ngaphambi kwexesha. I-EDTA ayiyi kufikelela kwisisombululo kuze kube yilapho i-pH ishintshwe malunga ne-8.0. Isisombululo se-500-milliliter yesitoreji se-0.5 M EDTA, isilinganise i-93.05 amagremu we-EDTA ityuwa ye-disodium (FW = 372.2). Emva koko uchithe kwi-400 milliliters amanzi adibeneyo kwaye utshintshe i-pH ne-NaOH. Emva koko, uphakamise isisombululo kwisikhululo sokugqibela sama-milliliters ama-500.
Lungisa iSolution Solution yeTBE
Yenza isisombululo esisisigxina (5x) sesitotyi se-TBE ngokulinganisa i-54 g ye-Tris base (FW = 121.14) kunye ne-27.5 g ye-boric acid (i-FW = 61.83) kwaye ichithe zombini kuma-milliliters angama-900 wamanzi achithwayo. Bese wongeza ii-milliliters ezingama-0.5 ze-0.5 M EDTA (pH 8.0) uze ulungise isisombululo kwisantya sokugqibela se 1 litre.
Esi sombululo sinokugcinwa kwindawo yeqondo lokushisa kodwa i-precipitate iya kubumba kwizisombululo ezindala. Gcina i-tampu kwiibhotile zeglasi uze ulawule ukuba kukho i-precipitate.
Lungiselela isisombululo sokuSebenza seTBE
I-agarose i-gel electrophoresis, i-TBE buffer ingasetyenziselwa ukuxinwa kwe-0.5x (1:10 ukukhutshwa kwe-stock). Nciphisa isisombululo se-stock ngo-10x kumanzi afakwe ngamanzi. Iziphumo zokugqibela ze-soliti zi-45 mM iTris-borate kunye ne-EDM eyi-1 yemitha. I-buffer manje ilungele ukusetshenziswa kwi -gel agarose .
Ingaba ufuna ntoni
Ukwenza i-TBE buffer, uzakufuna izinto ezine kuphela. Izinto ezisele kuloluhlu zixhobo. Izinto ezine ezifunekayo yi-EDTA ityuwa ye-saldium, isiseko seTris, i-boric acid kunye namanzi adibeneyo.
Ngokuphathelele izixhobo, uya kufuna imitha ye-pH kunye nemigangatho yokulinganisa, ngokufanelekileyo. Ukongeza koko, uya kufuna ama-600-milliliter kunye ne-1500-milliliter beaker okanye iiflasks. Ukugqitywa kwezixhobo zakho izixhobo ziphunyeziweyo kunye nezixhobo zokugubha kunye neeplate.
Hlola i-inventory kwibra oyisebenzisayo ukuqinisekisa ukuba unayo yonke into oyifunayo ngaphambi kokuba uqalise. Akukho nto yimbi ngaphezu kokuma phakathi kokulungiselela isisombululo ngenxa yokuba uphumelele kwizinto ezifanelekileyo.
Ukuba ibhanti yakho esikolweni okanye kwindawo yakho yomsebenzi, jonga abasebenzi abonekileyo ukuba babone yonke into esitokisini. Ukwenza oko kunokukugcina ixesha kunye namandla ekupheleni.