Enzymes nyinshi zikoreshwa mubushakashatsi bwa PCR

Urunigi rwa polymerase reaction, mu magambo ahinnye nkaPCRmucyongereza, ni tekinike ya biologiya ikoreshwa muguhuza ibice bya ADN byihariye.Irashobora gufatwa nka ADN idasanzwe yigana hanze yumubiri, ishobora kongera cyane ADN nkeya.Muri byosePCRuburyo bwo kubyitwaramo, icyiciro kimwe cyibintu bigira uruhare runini - enzymes.

1. Taq ADN

Mubigeragezo muminsi yambere yaPCR, abahanga bakoresheje Escherichia coli ADN polymerase ya I, Ariko hariho ikibazo cyiyi misemburo: igomba kuzuza enzyme nshya igihe cyose ikizamini cyakozwe, bigatuma intambwe yibikorwa igorana gato kandi biragoye guhita byiyongera.Iki kibazo cyakemutse nyuma y’uko abahanga batandukanije impanuka ya Taq ADN polymerase na Thermus aquaticus mu 1988. Kuva icyo gihe, kwongera kwa ADN mu buryo bwikora byabaye impamo.Ivumburwa ryiyi misemburo nayo irakoraPCRtekinoroji yoroshye, ifatika kandi ikoranabuhanga.Kugeza ubu, Taq ADN polymerase niyo polymerase ikunze kugaragara mubikoresho bya ADN.

2. PfuDNA

Nkuko byavuzwe haruguru, Taq DNase ifite ikosa rinini, bityo abahanga bahinduye polymerase ya Taq ADN ku rugero runaka kugirango birinde kwongerwaho bidasanzwe kubera kudahuza, bikavamo ibisubizo by’ibizamini bidahwitse.Ariko guhindura Taq polymerase ya Taq irashobora guhagarika ibikorwa bya ADN polymerase mubushyuhe bwicyumba.PfuDNA polymerase irashobora gukemura neza ibibi byavuzwe haruguru bya Taq ADN polymerase, kugirango PCR ishobore gukorwa mubisanzwe, kandi igipimo cyo gutsinda cya gene amplification gishobora kunozwa neza.

3. Hindura inyandiko mvugo

Reverse transcriptase yavumbuwe mu 1970. Iyi misemburo ikoresha RNA nk'icyitegererezo, dNTP nka substrate, ikurikiza ihame ryo guhuza ibice, kandi igahuza umurongo umwe wa ADN wuzuzanya na RNA mu cyerekezo cya 5′-3 ′.Guhindura inyandiko-mvugo ishingiye cyane cyane kubikorwa bya ADN polymerase biva muri ADN cyangwa RNA bityo ntigikorwa cya 3′-5 ′.Ariko, ifite ibikorwa bya RNase H, bigabanya synthesis uburebure bwa reverse transcriptase kurwego runaka.Bitewe n'ubudahemuka buke hamwe nubushuhe bwa transcriptque yinyamanswa, abahanga nabo barabihinduye.

KuriPCRubushakashatsi, ibyingenzi byingenzi ni tube umuyoboro wa PCR kugiti cye, 4/8-umurongo wa PCR, plaque ya PCR.

Labio'sIbikoresho bya PCRgira ibi bikurikiraibyiza:

Ibyapa bya PCR: Umuyoboro mugari wa cycler uhuza;Bitandukanye cyane, byoroshye kumenyekana neza;neza ibitekerezo bya fluorescence; byizaguhererekanya ubushyuhe;DNase yemewe, RNase, ADN, PCR inhibitor, kandi yapimwe pyrogen idafite.

Imiyoboro ya PCR kugiti cye: Imyuka irwanya umwuka; byizaguhererekanya ubushyuhe;Ibyiza bya optique bisobanutse; DNase yemewe, RNase, ADN, inhibitor za PCR, hamwe na pyrogene idafite.

4/8-imirongo ya PCR: Urukuta ruhebuje; rusobanutse neza; ibitekerezo byiza bya fluorescence;Irashobora gukoresha mu nganda zimiti, inganda zibiribwa, na biologiya ya biologiya; ibikoresho byiza cyane, ibikoresho bya PP;