zeziphi ii-organelles ezimbini ezingenakwahlulwa ngokulula nge-centrifugation
Ukwahlukana kweeseli ezingaphantsi: Ukoyisa uMngeni woKwahlula iiNuclei kunye neMitochondria
Intshayelelo:
I-Centrifugation yindlela enamandla esetyenziswa kwi-cell and molecular biology ukwahlula nokwahlula ii-organelles ezahlukeneyo kumxube ongafaniyo. Le nkqubo isekelwe kumgaqo we-differential sedimentation, apho ii-organelles ezinzima zihlala ezantsi kwetyhubhu phantsi kwamandla e-centrifugal. Nangona kunjalo, kukho izinto ezimbalwa ezingaqhelekanga kulo mgaqo, kwaye ii-organelles ezimbini ezingahlukaniyo ngokulula nge-centrifugation zii-nuclei kunye ne-mitochondria. Kweli nqaku, siza kuhlola imingeni ebandakanyekayo ekuhlukaniseni ezi organelles zimbini zibalulekileyo, ukubaluleka kokuhlanjululwa kwazo ngokubambisana kwiimvavanyo ezithile, kunye neendlela ezizezinye zokoyisa lo mqobo.
1. Ubunzima beNuclei kunye neMitochondria:
IiNuclei, ezidla ngokubizwa ngokuba ziziko lomyalelo weseli, zinoxanduva lokugcina izinto zemfuza kunye nokulawula ukubonakaliswa kwemfuza. Kwelinye icala, iiMitochondria ziindawo ezibalulekileyo ezivelisa amandla ngendlela ye-adenosine triphosphate (ATP). Ngenxa yemisebenzi yazo eyahlukileyo kunye neempawu zayo ezizodwa, ukwahlula ezi organelles zimbini kunokuba ngumceli mngeni omkhulu.
2. Ukubaluleka kokucoca kunye:
Nangona kunokuba yinto enqwenelekayo ukucoca ii-organelles nganye ukuze kwenziwe uvavanyo oluthile, kukho iimeko apho ukucocwa kwee-nuclei kunye ne-mitochondria kubaluleke kakhulu. Umzekelo, xa kufundwa ukulawulwa kokubonakaliswa kwezakhi zofuzo ukuphendula utshintsho kwi-metabolism ye-mitochondrial, kuba yinto ebalulekileyo ukuhlalutya impembelelo yokungasebenzi kakuhle kwe-mitochondrial ekubonakalisweni kwezakhi zofuzo zenyukliya. Kwiimeko ezinjalo, ukwahlukana kwee-nuclei kwi-mitochondria kuya kukhokelela kuhlalutyo olungaphelelanga kunye nokutolikwa okungachanekanga.
3. Imida ye-Centrifugation eqhelekileyo:
Iindlela zemveli zokucoca i-centrifugation zisebenzisa i-differential centrifugation, equka uthotho lwamanyathelo okucoca i-centrifugation ngesantya esahlukileyo ukuze kwahlulwe ii-organelles ngobukhulu nangobuninzi bazo. Ngelishwa, ii-nuclei kunye ne-mitochondria zinobunzima obufanayo kunye neempawu ezifanayo zokucoca i-sedimentation, okwenza kube nzima ukuzihlula ngokusebenzisa le ndlela yodwa. Oku kungenxa yomxholo ophezulu we-lipid kwii-organelles zombini, okukhokelela ekubeni zifane zijikeleze kwaye zibangele ukucoca i-co-sedimentation ngexesha lokucoca i-centrifugation.
4. Iindlela Ezizezinye Zokuqhelanisa:
Ukuze boyise imida ye-centrifugation, abaphandi baye baphuhlisa iindlela ezizezinye zokwahlula ii-nuclei kunye ne-mitochondria. Enye yeendlela ezinjalo yi-density gradient centrifugation, esebenzisa i-density gradient medium ukwenza i-gradient eqhubekayo. Ngokubeka i-homogenate ngononophelo phezu kwe-gradient kunye nokuyi-centrifuging, ii-organelles zinokuhlulwahlulwa ngokusekelwe kubunzulu bazo, zivumele ukwahlula ii-nuclei kunye ne-mitochondria ngendlela efanelekileyo. Nangona kunjalo, nangona ukwahlukana okuphuculweyo, le ndlela ingathatha ixesha elide kwaye kunzima ukuyenza.
5. Iindlela Eziphambili: IiBeads ZeMagnetic kunye neFluorescent Tagging:
Kwiminyaka yakutshanje, iindlela eziphambili ezibandakanya ii-magnetic beads kunye ne-fluorescent tagging ziye zathandwa kakhulu ekuhlukaniseni ii-subcellular. Ii-magnetic beads ezigqunywe ngee-antibodies ezithile ezijolise kwiiproteni zomphezulu ezikhoyo kwi-nuclei okanye kwi-mitochondria zingasetyenziswa ukwahlula i-organelle enye kwenye. Ngaphezu koko, ngokusebenzisa iidayi ezithile ze-fluorescent ezijoliswe kwi-nuclei okanye kwi-mitochondria, ii-organelles zinokuhlelwa kusetyenziswa ii-flow cytometers ezikhethekileyo okanye oomatshini be-fluorescence-activated cell sorting (FACS). Ezi teknoloji ziphambili zibonelela ngokuchaneka okungakaze kubonwe ngaphambili kunye nokusebenza kakuhle ekuhlukaniseni ezi organelles.
Isiphelo:
Ukwahlula ii-nuclei kunye ne-mitochondria ngumceli mngeni ngenxa yobuninzi bazo obufanayo kunye ne-lipid-rich. Ngelixa iindlela zendabuko zokucoca i-centrifugation zisokola ukufikelela ekuhlukaniseni okwaneleyo, iindlela eziphambili ezifana ne-density gradient centrifugation, i-magnetic bead-based isolation, kunye ne-fluorescent tagging ziye zavela njengezixhobo ezinamandla zokoyisa le miqobo. Ezi ndlela zintsha aziphuculi kuphela ukusebenza kakuhle kunye nokuchaneka kokwahlula i-organelle kodwa zikwavumela uhlalutyo olupheleleyo lwemisebenzi yazo kunye nokusebenzisana kwazo, zivula iindlela ezintsha zophando lweseli kunye ne-molecular.
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