Izinto ezininzi eziguqulweyo kwimbali yetekhnoloji yobhaqo engqondweni yam yitekhnoloji yokhuselo lomzimba olusekwe kumgaqo wokubophelela okukhethekileyo kwe-antigen-antibody, itekhnoloji ye-PCR kunye netekhnoloji yokulandelelana.Namhlanje siza kuthetha ngeteknoloji yePCR.Ngokutsho kokuvela kobuchwepheshe be-PCR, abantu bahlala behlula itekhnoloji ye-PCR ibe zizizukulwana ezithathu: itekhnoloji ye-PCR eqhelekileyo, itekhnoloji yexesha lokwenyani le-fluorescent yobungakanani be-PCR kunye netekhnoloji yedijithali yePCR.
Cubuchule bePCR obuqhelekileyo
UKARY MULLIS (1944.12.28-2019.8.7)
U-Kary Mullis waqulunqa i-polymerase chain reaction (i-polymerase chain reaction, i-PCR) ngo-1983. Kuthiwa xa wayeqhuba intombi yakhe, ngokukhawuleza waba ne-flash of inspiration kwaye wacinga ngomgaqo we-PCR (kwiinzuzo zokuqhuba).UKary Mullis wawongwa ngeBhaso likaNobel kwiChemistry ngowe-1993. IThe New York Times yagqabaza yathi: “Eyona nto iphambili nebalulekileyo, phantse iyahlulahlula ibhayoloji ibe ngamaxesha angaphambi kwePCR nasemva kwePCR.
Umgaqo we-PCR: Ngaphantsi kwe-catalysis ye-polymerase ye-DNA, i-DNA ye-strand yomama isetyenziswa njenge-template, kwaye i-primer ethile isetyenziswe njengendawo yokuqala yokwandisa, kunye nentombi ye-strand ye-DNA ehambelana ne-DNA ye-strand template yomama ikopishwa kwi-vitro ngokusebenzisa i-denaturation, i-annealing, ulwandiso kunye namanye amanyathelo.Yi-DNA synthesis amplification technology in vitro, enokuthi ngokukhawuleza kwaye ikhulise ngokuthe ngqo nayiphi na i-DNA ekujoliswe kuyo kwi-vitro.
Izinto ezilungileyo zePCR eqhelekileyo
1.Indlela yeClassic, imigangatho epheleleyo yamazwe ngamazwe kunye nekhaya
2.Iindleko eziphantsi zeerejenti zesixhobo
3.Iimveliso zePCR zinokufunyanwa ezinye iimfuniselo zebhayoloji yemolekyuli
Kucetyiswa umatshini we-Foregene PCR: https://www.foreivd.com/foreamp-sn-695-series-thermal-cycler-96-wells-pcr-machine-product/
Iimveliso ezinxulumeneyo: https://www.foreivd.com/pcr-herotm-with-dye-product/
Ukungalungi kwe-PCR eqhelekileyo
1.kulula ukungcolisa
2.ukusebenza nzima
3.kuphela uhlalutyo lobulunga
4.Uvakalelo oluphakathi
5.Kukho ulwandiso olungelulo olulodwa, kwaye xa ibhendi engayodwa ilingana nebhendi ekujoliswe kuyo, ayinakuhlukaniswa.
Ci-apillary electrophoresis-based PCR
Ukuphendula kwiintsilelo ze-PCR eziqhelekileyo, abanye abavelisi baye bazisa izixhobo ezisekelwe kumgaqo we-capillary electrophoresis.Isinyathelo se-electrophoresis emva kokukhulisa i-PCR kugqitywe kwi-capillary.Uvakalelo luphezulu, kwaye umahluko weziseko ezininzi unokuhlukaniswa kwaye ukukhulisa kunokubalwa ngu-MAERKER.umxholo wemveliso.Ukungalungi kukuba imveliso ye-PCR isafuna ukuvulwa kwaye ifakwe kwisixhobo, kwaye kusekho umngcipheko omkhulu wokungcola.
Ci-apillaryElectrophoresis
2. Ixesha lokwenyani le-fluorescent quantitative PCR (Quantitative Real-time PCR, qPCR) ubuchwephesheI-PCR ye-Fluorescent quantitative, ebizwa ngokuba yi-Real-Time PCR, iteknoloji entsha ye-nucleic acid quantitative eyaphuhliswa yi-PE (Perkin Elmer) ngo-1995. Imbali yophuhliso lwe-fluorescent quantitative PCR yimbali yomzabalazo ovuselela umphefumlo wezigebenga ezifana ne-ABI, i-Roche, kunye ne-Bio-Rad.Ukuba unomdla, ungayijonga.Obu buchule okwangoku bulolona buchule buqolileyo nobusetyenziswa ngokubanzi be-PCR obuphakathi.
Umatshini we-qPCR ocetyiswayo: https://www.foreivd.com/mini-real-time-pcr-system-forequant-sf2sf4-product/
Indlela yedayi yeFluorescent (SYBR Green I):I-SYBR Green I yeyona dayi isetyenziswa ngokuqhelekileyo kwi-DNA-binding ye-PCR yobungakanani, ebophelela ngokungangqalanga kwi-DNA ephindwe kabini.Kwimeko ekhululekileyo, i-SYBR Green ikhupha i-fluorescence ebuthathaka, kodwa xa ibotshelelwe kwi-DNA enemisonto ephindwe kabini, i-fluorescence yayo yonyuka ka-1000.Ke ngoko, isiginali ye-fluorescent iyonke ekhutshwe yi-reaction ilingana nobungakanani be-DNA ephindwe kabini ekhoyo kwaye iya kunyuka ngokunyuka kwemveliso ekhulisiwe.Ekubeni idayi ibophelela ngokungangqalanga kwi-DNA enemisonto ephindwe kabini, iziphumo ezilungileyo zobuxoki zinokuveliswa.
Iimveliso ezinxulumeneyo: https://www.foreivd.com/real-time-pcr-easytm-sybr-green-i-kit-product/
Indlela yokuhlola iFluorescent (iteknoloji yeTaqman): NgexeshaUkwandiswa kwe-PCR, i-probe ethile ye-fluorescent yongezwa ngexesha elifanayo njengesibini se-primers.I-probe yi-oligonucleotide yomgca, kunye neqela lentatheli ye-fluorescent kunye neqela le-fluorescent quencher ngokulandelanayo elibhalwe kuzo zombini iziphelo.Xa i-probe ihambelana, umqondiso we-fluorescent okhutshwe liqela leengxelo lixutywe liqela le-quencher, kunye nokufumanisa Akukho mqondiso we-fluorescent;ngexesha lokwandiswa kwe-PCR (kwinqanaba lokwandiswa), i-5'-3 'Dicer umsebenzi we-Taq enzyme iya kucola kwaye ithobe i-probe, ukuze iqela le-fluorescent lentatheli kunye neqela le-fluorescent le-quencher lihlulwe, ukwenzela ukuba inkqubo yokubeka iliso ye-fluorescence Isignali ye-fluorescent ingafunyanwa, oko kukuthi, rhoqo xa i-molecule ye-DNA iqonda i-chain chain ye-fluorescent i-synconcrescent ukuqokelelwa kwemiqondiso ye-fluorescent kunye nokwakhiwa kweemveliso ze-PCR.Indlela ye-Taqman probe yeyona ndlela ixhaphakileyo yokufumanisa ekubhaqweni kweklinikhi.
Iimveliso ezinxulumeneyo: https://www.foreivd.com/quickeasy%e1%b5%80%e1%b4%b9-real-time-pcr-kit-taqman-product/
Izinto ezilungileyo ze-qPCR
1.Indlela ivuthiwe kwaye izixhobo ezixhasayo kunye nee-reagents zigqityiwe
2.Iindleko eziphakathi zeerejenti
3.kulula ukuyisebenzisa
4.Ukuqonda okuphezulu kokuqonda kunye neenkcukacha
Ukungalungi kwe-qPCR
Utshintsho lwejini ekujoliswe kulo lukhokelela ekuphosweni kofuzo.
Isiphumo sokubhaqa se-low-concentration template asikwazi ukumiselwa.
Kukho impazamo enkulu xa usebenzisa igophe elisezantsi lobhaqo lomyinge.
3. I-PCR yedijithali (i-PCR yedijithali, i-dPCR) iteknoloji
I-PCR yeDijithali yindlela yokwenza umyinge opheleleyo we-nucleic acid.Xa kuthelekiswa ne-qPCR, i-PCR yedijithali inokufunda ngokuthe ngqo inani leemolekyuli ze-DNA/RNA, eyona milinganiselo ipheleleyo yeemolekyuli ze-nucleic acid kwisampulu yokuqala.Kwi-1999, uBert Vogelstein noKenneth W. Kin-zler bacebise ngokusemthethweni ingcamango ye-dPCR.
Ngo-2006, iFluidigm yaba yeyokuqala ukuvelisa isixhobo se-dPCR esekwe kwi-chip.Kwi-2009, i-Life Technologies yasungula inkqubo ye-OpenArray kunye ne-QuantStudio 12K Flex dPCR.Ngo-2013, i-Life Technologies yasungula inkqubo ye-QuantStudio 3DdPCR, esebenzisa i-high-density nanoscale microfluidic chip technology ukuze isasaze ngokulinganayo iisampuli kwiiseli ezingama-20,000.ekuphenduleni kakuhle.
Ngo-2011, i-Bio-Rad iqalise isixhobo se-droplet-based QX100 dPCR, esebenzisa iteknoloji yamanzi kwi-oyile ukuze isasaze ngokulinganayo isampuli kwi-20,000 ye-droplet-water-in-oil, kwaye isebenzisa i-droplet analyzer ukuhlalutya amaconsi.Kwi-2012, i-RainDance iqalise isixhobo se-RainDrop dPCR, eqhutywa yi-gas ephezulu yoxinzelelo, ukwahlula inkqubo nganye yokusabela kwi-emulsion yokusabela equkethe i-1 yezigidi ukuya kwi-10 yezigidi ze-picoliter-level micro-droplets.
Ukuza kuthi ga ngoku, i-PCR yedijithali yenze amaqela amabini amakhulu, uhlobo lwetshiphu kunye nodidi lwamathontsi.Kungakhathaliseki ukuba luhlobo luni lwe-PCR yedijithali, imigaqo yayo engundoqo iyanciphisa i-dilution, i-endpoint PCR kunye nokuhanjiswa kwe-Poisson.Inkqubo yokusabela ye-PCR eqhelekileyo equlethe itemplates ze-nucleic acid yahlulwe ngokulinganayo ibe amashumi amawaka eempendulo ze-PCR, ezisasazwa kwiichips okanye kwi-microdroplets, ukuze impendulo nganye iqulethe kangangoko kunokwenzeka i-molecule ye-template, kunye ne-molekyuli enye ye-PCR i-activation reaction yenziwa.Ngokufunda i-fluorescence Ubukho okanye ukungabikho komqondiso kubalwa, kwaye ubungakanani obupheleleyo benziwa emva kokulinganisa ukusabalalisa kwePoisson yezibalo.
Oku kulandelayo ziimpawu zamaqonga ePCR edijithali endikhe ndawasebenzisa:
1. Bio-Rad QX200 droplet digital PCR Bio-RadI-QX200 yi-platform ye-PCR ye-digital classic kakhulu, inkqubo yokufumanisa esisiseko: iisampuli ze-20,000 ziveliswa yi-droplet generator Amaconsi amancinci Amanzi-kwi-oyile akhuliswa kumatshini oqhelekileyo we-PCR, kwaye ekugqibeleni umqondiso we-fluorescence we-micro-droplet nganye ifundwa ngumfundi we-micro-droplet.Umsebenzi unzima ngakumbi, kwaye umngcipheko wongcoliseko uphakathi.
Xinyi TD1 micro-droplet yedijithali PCRI-Xinyi TD1 yiplatifomu ye-PCR yedijithali yasekhaya, inkqubo yokufumanisa eyisiseko: velisa amaconsi e-30,000-50,000 yamanzi kwi-oyile ngokusebenzisa i-droplet generator, ukukhulisa kwisixhobo esiqhelekileyo se-PCR, kwaye ekugqibeleni udlule Umfundi we-droplet ufunda isignali ye-fluorescent ye-droplet nganye.Zombini i-droplet generation kunye nokufunda kweli qonga zenziwa kwi-chip ezinikeleyo kunye nomngcipheko ophantsi wongcoliseko.
STILLA Naica micro-droplet chip yedijithali PCRSTILLA Naica liqonga elitsha ledijithali lePCR.Inkqubo yokufumanisa isiseko kukuba: yongeza isisombululo sokusabela kwi-chip, faka i-chip kwisizukulwana se-micro-droplet kunye ne-amplification system, kwaye uvelise ama-30,000 amaconsi amancinci.Ukusabalalisa kwi-chip, kwaye ukukhulisa i-PCR kugqityiwe kwi-chip.Emva koko i-chip ekhulisiwe idluliselwa kwinkqubo yohlalutyo lokufunda i-micro-droplet, kwaye isignali ye-fluorescent ifundwa ngokuthatha imifanekiso.Ekubeni yonke inkqubo yenzeke kwi-chip evaliweyo, umngcipheko wokungcola uphantsi.
4. ThermoFisher QuantStudio 3D chip yedijithali PCR
I-ThermoFisher QuantStudio 3D lelinye iqonga lePCR esekwe kwitshiphu yedijithali.Inkqubo yokufumanisa isiseko kukuba: yongeza isisombululo sokusabela kwi-spreader, kwaye usasaze isisombululo sokuphendula ngokulinganayo kwi-chip kunye ne-20,000 microwells ngokusebenzisa i-spreader., faka i-chip kumatshini we-PCR ukuze ukhulise, kwaye ekugqibeleni ubeke i-chip kumfundi kwaye uthathe ifoto ukuze ufunde isignali ye-fluorescent.Umsebenzi unzima kakhulu, kwaye yonke inkqubo iqhutyelwa kwi-chip evaliweyo, kwaye umngcipheko wokungcola uphantsi.
5. JN MEDSYS Ukucaca chip yedijithali PCR
I-JN MEDSYS Ukucaca liqonga elitsha le-PCR ledijithali.Inkqubo yokufumanisa isiseko kukuba: yongeza isisombululo sokuphendula kwi-applicator, kwaye usasaze isisombululo sokusabela ngokulinganayo kwiityhubhu ze-PCR eziyi-10,000 ezigxininiswe kwi-tube ye-PCR ngokusebenzisa umfaki-sicelo.Kwi-chip ye-microporous, isisombululo sokuphendula singena kwi-chip ngokusebenzisa i-capillary action, kwaye ityhubhu ye-PCR ene-chip ibekwe kumatshini we-PCR ukwenzela ukukhulisa, kwaye ekugqibeleni i-chip ifakwe kumfundi ukuba afunde umqondiso we-fluorescent ngokuthatha ifoto.Utyando lunzima ngakumbi.Umngcipheko wosulelo uphantsi.
Iiparamitha zeqonga ngalinye ledijithali lePCR zishwankathelwa ngolu hlobo lulandelayo:
Iimpawu zovavanyo lweqonga le-PCR yedijithali zezi: inani leeyunithi zokwahlula, inani leziteshi ze-fluorescent, ubunzima bokusebenza kunye nomngcipheko wokungcola.Kodwa eyona nto ibalulekileyo kukufumanisa ukuchaneka.Enye indlela yokuvavanya iiplatifti zedijithali zePCR kukusebenzisa iiplatifti ezininzi zedijithali zePCR ukuqinisekisana, kwaye enye indlela kukusebenzisa izinto ezisemgangathweni ezinexabiso elichanekileyo.
Izinto ezilungileyo zedPCR
1.Ukufezekisa ubungakanani obupheleleyo
2.Uvakalelo oluphezulu kunye neenkcukacha
3.Iyakwazi ukubona iisampuli zeekopi eziphantsi
Ukungalungi kwe-dPCR1. Izixhobo ezixabisa kakhulu kunye nee-reagents 2. Ukusebenza okuntsokothileyo kunye nexesha elide lokubona 3. Uluhlu lokufumanisa olumxinwa
Okwangoku, izizukulwana ezithathu zeteknoloji ye-PCR zineenzuzo zabo kunye nokungahambi kakuhle, kwaye nganye inemimandla yayo yesicelo, kwaye akusiyo ubudlelwane ukuba esinye isizukulwana sithatha indawo yesinye.Ukuqhubela phambili kwetekhnoloji kuye kwafaka amandla amatsha kwitekhnoloji yePCR, iyenza ukuba ikwazi ukuvula isalathiso sesicelo esinye emva kwenye, ukwenza ubhaqo lwe-nucleic acid lube lula kwaye luchaneke.
Umthombo: UGqr. Yuan uyakuthatha ayokuvavanywa
Iimveliso eziCetyisiweyo:
Ixesha lokuposa: Nov-18-2022
