Isolating DNA daga E. Coli wani tsari ne na asali a cikin ilimin kwayoyin halitta. Wannan talifin zai yi muku tafiya cikin dukkan aikin, yana samar da cikakken matakan da bayani, tabbatar muku fahimtar dukiyar da fannoni masu amfani. Ko dai mai bincike ne na roba ko sabon bincike ga lab, wannan jagorar zai zama hanya mai mahimmanci.
Shiri na dakatarwar sel
Tarin E. Cori Coli
Mataki na farko a cikin ware DNA daga E. Chi ya ƙunshi tattara ƙwayoyin ƙwayoyin cuta. Wannan yawanci yana buƙatar haɓakawa E. Chi Coli a cikin matsakaici mai dacewa har sai ya kai ga ci gaban logarithmic ci gaba. Lokaci yana da mahimmanci saboda sel a wannan lokaci shine mai yiwuwa kuma mafi sauƙin lyse, wanda zai haifar da yawan amfanin ƙasa.
● sel a cikin sandar da ta dace
An tattara ƙwayoyin ƙwayoyin da aka tattara sannan a cikin buffer da suka dace. Zabi na yau da kullun shine Tris - Edta (Te) Buffer, wanda ke taimaka wa amincin DNA yayin hakar. Buffer ya yi amfani da dalilai da yawa: Yana tabbatar da cations na FLALS wanda ba zai iya kashe DNA ba, kuma yana ba da ingantacciyar yanayin halayen enzymatic.
Centrifugation zuwa sel na pellet
● sigogi don centrifugation (saurin da lokaci)
Bayan ya sake yin sel, an dakatar da dakatarwar zuwa centrifugation zuwa pelllet da sel. Saurin hanzari da lokacin sigogi ne masu mahimmanci. Yawanci, ana yin centrifugation a kusa da 4,000 - 6,000 g don 10 - mintina 15 a 4 ° C. Wannan yana tabbatar da cewa sel samar da ƙarfi pellet a kasan bututun na centrifuge.
● Hasri mai mahimmanci game da kyakkyawan pelle
Pelsarfin da ya dace yana da mahimmanci don rarrabe ƙwayoyin daga matsakaici da sauran kayan haɗin mai narkewa. Da kyau - An kafa Pellet da matakai masu zuwa kuma mafi inganci, saboda haka, yawan yawan amfanin ƙasa.
Cire SuperNat
● dabaru don cirewa
Da zarar an fifita sel, masu suttura (na ruwa a sama da pellet) dole ne a cire su ba tare da tayar da pellet ɗin tantanin halitta ba. Ana yin wannan yawanci ta amfani da micropipette. Yana da mahimmanci don aiwatar da wannan matakin sosai don guje wa rasa kowane sel.
● tabbatar da asarar karancin tantanin halitta
Tabbatar da asarar karar tantanin halitta ya ƙunshi bututun ƙarfe kuma, idan ya cancanta, da yawa na cirewar centrifugh da kuma sananniyar central. Manufar shine a ci gaba da kwayoyi da yawa kamar yadda zai yiwu a cikin pellet don iyakar dawo da DNA.
Bugu da kari na nuclei lysis bayani bayani
● Kayayyakinan unseran na uclei lysis
Kungiyar kare heplei na yawanci tana dauke da kayan maye (kamar sds), buffer (kamar tris - hcl), da kuma wakili mai kauri (kamar Edta). Jirgin ruwan wanka da ambulaf na nukiliya, suna sakin abin da ke ciki, gami da DNA, cikin Magani.
● rawa a cikin rushe bangon tantanin halitta
Finalin nuclei na kare ba kawai ya lalata ƙwayar tantanin halitta ba har ma da na deNaurizes sunadarai da lipids bango da kuma ambulan makaman tantanin halitta don saki DNA zuwa cikin mafita.
Resusepevenvent na sel
Mory Pipeting buteting don kauce wa DNA shearing
Da zarar an ƙara maganin nuclei lyisi, ana buƙatar sel suna buƙatar sake komawa a hankali don gujewa DNA. Shearing na iya karya DNA zuwa ƙananan gutsuttsura, waɗanda na iya zama matsala ga aikace-aikacen ƙasa waɗanda ke buƙatar High - Motar nan nauyi -
● tabbatar da cikakken gyara
Cikakken jerin abubuwan da ya tabbatar da cewa duk sel suna kwance gabaɗaya, haɓaka maɓallin dawowar DNA. Ana iya samun wannan ta hanyar pipetting mai laushi ko vortexing mafita a ƙarancin gudu.
Shiryawa ga sel Lyse
● Saitunan zazzabi don shiryawa
Kwayoyin sel na kwafa a cikin yanayin zazzabi don tabbatar da cikakken lysis. Ana yin wannan yawanci a 37 ° C zuwa 55 ° C. Ainihin zazzabi da tsawon lokaci na iya bambanta dangane da yarjejeniya da takamaiman buƙatun na kayan aikin DNA suna amfani da su.
Harkokin da ake buƙata don ingantaccen lysis
Tsawon lokaci na shiryawa shine tsakanin minti 20 zuwa 30, amma ana iya gyara shi dangane da ingancin ilimin tantanin halitta lura. Tsawo bayani na iya zama dole don cikakken lysis amma ya kamata a daidaita kan haɗarin lalata DNA.
Sanyaya zuwa zazzabi a daki
● Muhimmancin sanyaya hankali
Bayan shiryawa, a hankali ana sanyaya wa zafin jiki a cikin zazzabi. Sanyaya sanyaya yana taimakawa wajen inganta DNA kuma yana rage haɗarin girgizar zafi, wanda zai iya rage DNA.
● Sakamakon akan DNA da kuma tarkace
Sanyaya zuwa zazzabi ɗaki yana ba da dafar tarkace ta salula don yin hazo, yana sauƙaƙa raba DNA a matakai masu zuwa. Wannan kuma yana taimakawa wajen aiwatar da ayyukan enzyme kuma yana sauƙaƙe cire RNA by Rnase magani.
Addarin bayani rnase bayani
Man NOMAN DUKA KYAUTA A CIKIN SAUKI
An kara da maganin Rnase a Degrade RNA, wanda zai iya inna a co - tsarkake tare da DNA da kuma tsoma baki tare da aikace-aikacen ƙasa. Rnase zabi tsintsaye RNA, barin DNA m.
● Ana hana gurbun RNA
Yana hana gurbata RNA yana da mahimmanci ga aikace-aikace waɗanda ke buƙatar DNA tsarkakakkiyar DNA, kamar PCR da Searquin. Jiyya na RNase yana tabbatar da cewa an ware DNA na ware daga RNA mashedawa.
Tsarkake DNA
● Hanyoyi don raba DNA daga wasu kayan aikin salula
Ana iya amfani da hanyoyi da yawa don tsarkaka da DNA daga Lysate. Waɗannan sun haɗa da phenol - hakar chloroform, ethanol hazo, da kuma amfani da kasuwanci E. Cori Dna. Kowace hanya tana da ribobi da fursunoni, tare da kayan kasuwanci suna ba da dacewa da daidaitaccen sakamako.
● tunanin da DNA tsarkakewa
Tsarkake mai matukar muhimmanci ga nasarar aikace-aikacen ƙasa. Kasuwanci, kamar su tantanin halitta na E. Cori DNA Kit, an tsara su ne don samar da manyan - Tsarkakakken lipids, da sauran manyan gurbata.
Ajiya da sarrafa dNA
● mafi kyawun ayyukan don adana DNA
Da zarar an tsarkaka, ya kamata a adana DNA a cikin Buffer da suka dace, kamar Sipper, a - 20 ° C ko - Adadin 80 ° C na dogon lokaci. Guji daskare akai-akai - Thaw hycles kamar yadda suke iya haifar da lalata DNA.
● Kula da amincin DNA don Aikace-aikacen ƙasa
Don kula da amincin DNA, Yi amfani da bakararre bakararre, nuclease - tubes kyauta da mafita. Wannan yana tabbatar da cewa DNA ya rage kuma ya dace don aikace-aikace kamar cloning, sequincing, da PCR.
KayiAlishir
Jiangsu Hillsugene ya kafa hedkwatar ta (tsire-tsire 10,000㎡) da kuma cibiyar R & Dozong) a cikin Shenzhen da Shanghore guda biyu a Shenzhen da Shanghai, wanda ke shimfida hanyar sadarwa guda biyu a kasar. Saƙon soja na Arewa a Amurka a yanzu yana kan gini, yana ƙara yada kasancewar ta duniya. Hillgene ya gina hanyar musayar hanya don haɓaka samfuran warkarwa, daga gano kayan aikin masana'antu na acid, Magani - dakatarwar tsari, an dakatar da tsarin aiwatarwa, da gwajin tsarin, da gwajin aiwatarwa. An tsara samfuran Bluekekit don ikon sarrafa ingancin, tabbatar da nasarar sabbin kayan salula.
Lokaci: 2024 - 05 - 05 14:47:03
