Kumisikidzwa kwemaitiro kunosanganisira optimization yavo uye kunotevera kumisikidzwa uye kuwiriranisa, zvichibvumira kwenguva refu yakakwana kuita - yakazvimirira pamushandisi. Kumira kunovimbisa mhedzisiro yemhando yepamusoro, pamwe nekuberekana kwavo uye kuenzaniswa.
Chinangwa che (classic) PCR chizvarwa chezvakavimbika uye zvinogoneka. Kune mamwe maapplication, goho rePCR chigadzirwazvine basa zvakare. Nezvekuita uku, kutarisirwa kunofanirwa kutorwa kuti ive nechokwadi chekuti masampuli haana kukanganisa uye kuti PCR workflow inoramba yakagadzikana. Kunyanya, izvi zvinoturikira kudzikisira kuunzwa kwekusvibiswa kunogona kutungamira kune yenhema yakanaka kana yenhema mhedzisiro yakaipa kana kuvharisa maitiro ePCR. Uyezve, maitiro ekuita anofanirwa kunge akafanana sezvinobvira kune yega yega sampu mukati mekumhanya uye zvakare kuchinjirwa kune anotevera maitiro (enzira imwe chete). Izvi zvinoreva kuumbwa kwemaitiro pamwe chete nemhando yekudzora tembiricha mubhasikoro. Zvikanganiso zvemushandisi, hongu, zvinofanirwa kudziviswa zvakanyanya sezvinobvira.
Pazasi, isu ticharatidza matambudziko anowanikwa panguva yekugadzirira uye panguva yese yePCR - uye nzira dzemhinduro dziripo maererano nezviridzwa uye zvinodyiwa zvinoshandiswa pakumisikidza kwePCR workflows.
Kugadzirira kuita
Kugovera kwezvinoita zvinhu muPCR-midziyo, kana mahwendefa, zvichiteerana, kunosanganisira matambudziko akawanda anofanirwa kukundwa:
Maitiro ekuita
Iyo chaiyo uye chaiyo dosing yezvikamu zvega zvega zvakakosha kana uchivavarira mamiriro akafanana ekuita anogoneka. Pamusoro peiyo yakanaka pipetting tekinoroji, kusarudza chishandiso chakakodzera kwakakosha. PCR master-mix inowanzove ine zvinhu zvinowedzera viscosity kana kuburitsa furo. Munguva yepipetting process, izvi zvinotungamira mukunyorova kukuru kweiyopipette mazano, nokudaro kuderedza pipetting kururama. Kushandiswa kwemaitiro ekuparadzira zvakananga kana mamwe mazano epipette asinganyanyi kunyorova anogona kuvandudza kurongeka uye kurongeka kwepipetting process.
Contaminations
Munguva yekugovera, maaerosols anogadzirwa ayo, kana akabvumidzwa kusvika mukati mepipette, anogona kusvibisa imwe sampuli panguva inotevera yepipetting nhanho. Izvi zvinogona kudzivirirwa nekushandisa matipi esefa kana masisitimu ekutamisa akananga.
Consumables akadaimazano, midziyo uye mahwendefa anoshandiswa muPCR workflow haifanire kunge iine zvinhu zvinokanganisa sampuli kana manyepo mhedzisiro. Izvi zvinosanganisira DNA, DNases, RNases uye PCR inhibitors, pamwe chete nezvikamu zvingango bvisa kubva muzvinhu panguva yekuita - zvinhu zvinozivikanwa sezvinobviswa.
Kukanganisa kwemushandisi
Iyo yakawanda sampuli inogadziriswa, inowedzera kukanganisa kukanganisa. Zvinogona kuitika nyore kuti sampuli inopinzwa mumudziyo usiri iwo kana tsime risiri iro. Ngozi iyi inogona kudzikiswa zvakanyanya nekuisa chiratidzo mumatsime zviri nyore. Kuburikidza neautomation yematanho ekuparadzira, iyo "human factor", kureva, kukanganisa uye misiyano inoenderana nemushandisi, inoderedzwa, nekudaro ichiwedzera kuberekana, kunyanya mune madiki maitiro mavhoriyamu. Izvi zvinoda ma plates ekudzikama kwakaringana kuti ashandiswe munzvimbo yekushandira. Mabhakodhi akabatanidzwa anopa kuwedzera kuverenga-muchina, iyo inorerutsa sampuli yekutevera mukati mese maitiro.
Kugadziriswa kweiyo thermocycler
Kugadzira chiridzwa chinogona kuratidza kuti chinotora nguva uye chinokanganisa. Yakasiyana PCR yekupisa cycler maficha anoshanda pamwechete kurerutsa iyi nhanho uye, zvakanyanya kukosha, kuti ive yakachengeteka:
Kushanda kuri nyore uye kutungamira kwakanaka kwemushandisi ndiyo hwaro hwehurongwa hunoshanda. Kuvaka panheyo iyi, password-yakachengetedzwa mushandisi manejimendi inodzivirira yako pachako zvirongwa kubva kuchinjwa nevamwe vashandisi. Kana akawanda macyclers (emhando imwechete) ari kushandiswa, zvinobatsira kana chirongwa chinogona kutamiswa zvakananga kubva kune chimwe chiridzwa kuenda kune chimwe kuburikidza ne USB kana yekubatanidza. Kombuta software inogonesa yepakati uye yakachengeteka kutonga kwezvirongwa, kodzero dzevashandisi uye zvinyorwa pakombuta.
PCR kumhanya
Munguva yekumhanya, DNA inokwidziridzwa mumudziyo wekuita, uko sampu yega yega inofanirwa kuiswa kune yakafanana, inopindirana maitiro ekuita. Aya anotevera maitiro akakosha kune iyo process:
Kudzora tembiricha
Kurongeka kwakanakisa mukudzora tembiricha uye homogeneity ye cycler block ndiyo hwaro hweiyo tembiricha yekumisikidza yemasampuli ese. Hunhu hwepamusoro hwekudziya uye kutonhora zvinhu (peltier elements), pamwe chete nenzira iyo izvi zvakabatana nayo kubhuroka, ndizvo zvinosarudza zvinosarudza njodzi yekusiyana kwekushisa kunozivikanwa se "edge effect"
Evaporation
Izvo zvakadzika zvezvimiro zvega zvega hazvifanirwe kuchinja pamusoro pekufamba kwekuita nekuda kwekubuda. Zvikasadaro, zvinogoneka kuti zvishomaPCR chigadzirwainogona kugadzirwa, kana kwete zvachose. Saka zvakakosha kuti uderedze kubuda kwemhepo nekusimbisa chisimbiso chakachengeteka. Muchiitiko ichi, chivharo chinopisa che thermocycler uye chisimbiso chemudziyo chinoshanda nemaoko. Mhando dzakasiyana dzekuisa chisimbiso dziripoPCR mahwendefa (chinongedzo: Chisimbiso chinyorwa), umo chisimbiso chakanakisisa chinowanikwa kuburikidza nekupisa kupisa. Kumwe kuvharwa kunogonawo kuve kwakakodzera, chero bedzi kudzvinyirirwa kwechivharo che cycler kunogona kugadziriswa kune chisimbiso chakasarudzwa.
Maitiro ekumira ari munzvimbo kuitira kuchengetedza zvakarurama uye zvinogoneka mhedzisiro munguva refu. Izvi zvinosanganisira kugadzirisa nguva dzose kwemichina kuti ive nechokwadi chokuti inogara iri mumamiriro akanaka ekushanda. Zvese zvinoshandiswa zvinofanirwa kunge zviri zvemhando yepamusoro pamijenya yese inogadzirwa, uye kuwanikwa kwavo kwakavimbika kunofanirwa kuvimbiswa.
Nguva yekutumira: Nov-29-2022