inoita kuti centrifuge iparadzanise DNA nemapuroteni

Ko Centrifuge inoparadzanisa DNA nemapuroteni here?

Nhanganyaya

Kubatanidza masero (centrifugation) inzira inokosha inoshandiswa zvakanyanya mukutsvagisa kwesainzi uye mumarabhoritari ekiriniki pakuparadzanisa zvikamu zvakasiyana zvemuenzaniso zvichienderana nehuwandu hwazvo. Imwe nzira inowanzo shandiswa yeiyi nzira inosanganisira kuparadzanisa DNA nemapuroteni kubva mumuenzaniso webhayoloji. Muchinyorwa chino, tichaongorora kana centrifuge ichikwanisa kuparadzanisa DNA nemapuroteni zvinobudirira uye toongorora nzira dzakasiyana dzinoshandiswa kuti tiwane kupatsanurana kwakanaka.

Kunzwisisa Centrifugation

Usati wanyatsoongorora magadzirirwo ekuparadzanisa DNA kubva kumapuroteni uchishandisa centrifuge, zvakakosha kunzwisisa misimboti yekutanga ye centrifuge. Centrifuge inoshanda pamisimboti ye sedimentation uye inoshandisa high-speed spinning kugadzira simba rakasimba re centrifugal. Simba iri rinoita kuti zvidimbu zviri mumuenzaniso zviparadzane zvichienderana nehuwandu hwazvo, nezvidimbu zvakakora zvichifamba zvichienda pasi pechubhu yemuenzaniso. Nekuda kweizvozvo, zvidimbu zvakareruka zvinowanzogara pamusoro.

Dambudziko Rekuparadzanisa DNA Nemapuroteni

Kana zvasvika pakuparadzanisa DNA nemapuroteni, zvinopa dambudziko sezvo mamorekuru ese ari maviri aine huremu hwakafanana hwemamorekuru. Pasina nzira inoshanda yekuparadzanisa, zvakaoma kuwana DNA chaiyo kana mapuroteni. Zvisinei, centrifugation inogona kubatsira mukuparadzanisa, asi inogona kusakwana yega.

1. Kusiyanisa Kuparadzanisa: Kuparadzana Kwekutanga

2. Ultracentrifugation: Kuparadzaniswa Kwakawedzerwa

3. Kudzora Kuwanda Kwemagiraidhi: Kugadzirisa Maitiro

4. Kukosha kweKusarudza Buffer

5. Kubatanidza simbi muchina uchienzaniswa nedzimwe nzira dzekuparadzanisa

Kuparadzanisa Kwepakati: Kuparadzana Kwekutanga

Kuparadzanisa DNA kubva kumapuroteni nenzira dzakasiyana ndiyo nhanho yekutanga mukuparadzanisa DNA kubva kumapuroteni. Nzira iyi inoshandisa kumhanya kwakasiyana kwekubatanidza DNA kuti iwane kupatsanurwa kwakakura kwezvikamu zvemasero. Munguva iyi, masero akafanana kana kuti sampuli dzematishu dzinoiswa muchikamu chemagetsi chinokasira kupatsanurwa, kazhinji kacho kanosvika 1000-5000 revolutions paminiti (rpm). Kutenderera uku kwekukurumidza kunogonesa kupatsanurwa kwemasero enuclei neasina kusimba, zvichikonzera kupatsanurwa kwemasero. Zvisinei, hazvipe kupatsanurwa kwakakwana kweDNA nemapuroteni.

Kupinza masero zvakanyanya: Kuparadzanisa Kwakawedzerwa

Kuti zvikunde zvipingamupinyi zve differential centrifugation, ultracentrifugation inopinda munzvimbo iyi. Ultracentrifuges ihigh-speed centrifuges inokwanisa kusvika kumhanya kunosvika 100,000 rpm. Nekuisa crude fraction inowanikwa kubva pakutanga low-speed centrifugation kuenda ultracentrifugation, huwandu hwakakwira hwekuparadzanisa DNA nemapuroteni hunogona kuwanikwa. Mamorekuru eDNA anorema anodonhera pasi pechubhu, achigadzira pellet inoonekwa, nepo mapuroteni achiramba ari musupernatant.

Kudzora Kuwanda Kwemagiraidhi Ekuwedzera: Kugadzirisa Maitiro

Kuti DNA nemapuroteni zvinyatso patsanurwa, kushandiswa kwe density gradient centrifugation. Munzira iyi, nzira ye density gradient medium, yakaita sesucrose kana cesium chloride, inogadzirwa. Iyi nzira ine huwandu hwakasiyana hwe densities, ichiwedzera kubva kumusoro kusvika pasi pechubhu ye centrifuge. Chikamu chisina kuchena chakawanikwa kubva padanho rekare chinoiswa pamusoro pe density gradient uye chinoiswa ultracentrifugation zvakare. DNA nemapuroteni zvinopatsanurwa zvichienderana nehuwandu hwazvo uye zvinoumba mabhendi akasiyana mukati me gradient. Kupatsanurwa kwe gradient kunobvumira kudzoserwa kweDNA yakachena nemapuroteni samples.

Kukosha kweKusarudza Buffer

Buffer inoshandiswa panguva ye centrifugation inoita basa rakakosha mukuwana kupatsanurwa kwakanaka. Inokanganisa kugadzikana uye kuvimbika kwemamorekuru ari kupatsanurwa. Kune DNA, Tris-EDTA buffer ine pH ye8.0 inowanzo shandiswa nekuda kwekukwanisa kwayo kuchengetedza kugadzikana kweDNA. Kune rumwe rutivi, mapuroteni anoda buffers ane pH levels chaiyo, kazhinji inenge 7.0, kudzivirira denaturation uye kuchengetedza chimiro chayo chepakutanga. Kusarudza buffer yakakodzera kunovimbisa kupatsanurwa kwakakodzera kweDNA nemapuroteni pasina kukanganisa kunaka kwazvo.

Kuparadzanisa midziyo yemagetsi (Centrifugation) kana kuti dzimwe nzira dzekuparadzanisa

Kunyange zvazvo centrifugation iri nzira inoshandiswa zvakanyanya mukuparadzanisa DNA kubva kumapuroteni, zvakakosha kubvuma kuti ingasava iyo chete nzira inoshandiswa. Dzimwe nzira, dzakadai se chromatography, electrophoresis, uye precipitation, dzinogona kuwedzera centrifugation kuti pave nekupatsanurwa kwakanyatsonaka. Nzira yega yega ine zvayakanakira nezvayakaipira, uye vaongorori vanowanzo sanganisa nzira dzakawanda kuti vawane kuchena uye kudzoreredzwa kweDNA nemapuroteni.

Mhedziso

Mukupedzisa, kunyange zvazvo centrifuge ichinyatsobatsira pakuparadzanisa DNA kubva kumapuroteni, inoshanda zvakanaka pamwe chete nedzimwe nzira. Kuparadzanisa centrifugation, ultracentrifugation, uye density gradient centrifugation zvinoumba matanho ekuparadzanisa anobatsira kuwana sampuro dzakachena. Pamusoro pezvo, kusarudza mabuffer akakodzera uye kufunga nezvedzimwe nzira dzekuparadzanisa zvinowedzera kushanda zvakanaka kwekuparadzanisa DNA nemapuroteni.