COVID-19 cuta ce mai saurin kamuwa da Cutar Cutar Cutar Coronavirus Nau'in 2. Lokacin da mutum ya kamu da cutar, alamomin da aka fi sani da su sun hada da zazzabi, tari, da ƙarancin numfashi.

Ana iya tattara samfuran da aka yi amfani da su don gwaji ta hanyar swabs na nasopharyngeal ko swabs na oropharyngeal.

Menene PCR?

Madaidaicin hanyar gano coronavirus shine maganin sarkar polymerase, PCR.Wannan hanya ce da ake amfani da ita sosai a ilmin halitta.Yana iya kwafin miliyoyi da sauri zuwa biliyoyin takamaiman guntu DNA.

Sabuwar coronavirus ta ƙunshi kwayar halittar RNA mai tsayi mai tsayi.Don gano waɗannan ƙwayoyin cuta ta PCR, ƙwayoyin RNA dole ne a canza su zuwa jerin DNA ɗin su na ƙarin ta hanyar jujjuya rubutun, sa'an nan kuma za a iya haɓaka sabon DNA ɗin ta daidaitattun hanyoyin PCR, wanda aka fi sani da RT-PCR.

RT-PCR tsari

RNA cirewa

Don aiwatar da wannan hanyar, dole ne a fitar da kwayar cutar RNA da gaske.Ana iya amfani da nau'ikan kayan aikin tsarkakewa na RNA don dacewa, sauri da kuma rabuwa mai tasiri.

Don cire kwayar cutar RNA ta amfani da kayan kasuwanci, da farko ƙara samfurin zuwa bututun microcentrifuge sannan a haɗa shi da ma'aunin lysis.Wannan buffer ɗin yana da ƙima sosai kuma yawanci ya ƙunshi phenol da guanidine isothiocyanate.Bugu da kari, masu hana RNase galibi suna kasancewa a cikin buffer lysis don tabbatar da keɓewar RNA mara lafiya.

Bayan ƙara lysis buffer, jujjuya bututun gauraya ta bugun bugun jini kuma a sanya shi a zafin jiki.Sannan ana lysed kwayar cutar a ƙarƙashin yanayin rashin ƙarfi sosai wanda ma'aunin lysis ya samar.

Bayan samfurin an lysed, ana amfani da bututu na centrifuge don aikin tsarkakewa.Ana ɗora samfurin a cikin bututun centrifuge sannan a ɗaure.

Wannan hanya hanya ce mai ƙarfi ta hakar lokaci wacce lokacin tsayawa ya ƙunshi matrix gel silica.

Ƙarƙashin mafi kyawun gishiri da yanayin pH, ƙwayoyin RNA suna ɗaure ga membrane na silica.

A lokaci guda kuma, ana cire furotin da sauran gurɓatattun abubuwa.

Bayan centrifugation, sanya bututun centrifuge a cikin bututu mai tsabta mai tsabta, jefar da tacewa, sa'an nan kuma ƙara buffer na wankewa.

Saka bututu a cikin centrifuge kuma don tilasta buffer ɗin wanke ta cikin membrane.Wannan zai cire duk sauran ƙazanta daga membrane, barin kawai RNA da ke ɗaure zuwa gel silica.

Bayan an wanke samfurin, sanya bututun a cikin bututun microcentrifuge mai tsabta kuma ƙara mai buffer elution.

Sa'an nan kuma an centrifud don tilasta ma'ajiyar elution ta cikin membrane.Buffer na elution yana cire RNA mai hoto hoto ko bidiyo mai zagaya yanar gizo da sauri kuma yana samun RNA mai tsafta ba tare da sunadarai, masu hanawa, da sauran gurɓatattun abubuwa ba.

MATAKI NA 2

Mixed maida hankali

Bayan fitar da kwayar cutar RNA, mataki na gaba shine a shirya cakudawar amsa don haɓaka PCR.A wannan mataki, ana amfani da hankali.Wannan bayani da aka tattara shine ingantaccen bayani mai tattara bayanai wanda ya ƙunshi premix, baya transcriptase, nucleotides, firamare na gaba, juzu'i, binciken TaqMan da DNA polymerase.

A ƙarshe, don kammala wannan cakudawar, ana ƙara samfurin RNA.Ana haxa bututun ta hanyar bugun bugun jini, sannan ana ɗora cakuɗen amsawa cikin farantin PCR.Farantin PCR yawanci yana ƙunshe da rijiyoyi 96 kuma yana iya nazarin samfurori da yawa a lokaci guda.

MATAKI NA 3

PCR haɓakawa

Bayan haka, sanya farantin a cikin injin PCR, wanda shine ainihin mai hawan zafi.

Ana amfani da RT-PCR na ainihi don gano sabon coronavirus na 2019 ta hanyar haɓaka jerin abubuwan da aka yi niyya a cikin jinsin RdrRP, E gene da N gene.Zaɓin nau'in halittar da aka yi niyya ya dogara da jerin abubuwan farko da bincike.

Mataki na farko na RT-PCR shine juyar da rubutu.Sashin farko na ƙarin DNA an haɗa shi, wanda PCR reverse primer ya qaddamar, wanda ke ɗaure ga ɓangaren madaidaicin kwayar halitta ta RNA.Sa'an nan reverse transcriptase yana ƙara DNA nucleotides zuwa 3'ƙarshen farko na farko don haɗa DNA mai dacewa da RNA mai hoto mai hoto.Matsakaicin zafin jiki da tsawon lokacin wannan matakin sun dogara da maƙallan maɓalli, RNA manufa, da juyar da rubutun da aka yi amfani da su.

Bayan haka, ana amfani da matakin denaturation na farko, wanda ke haifar da raguwar matasan RNA-DNA.Wannan mataki ya zama dole don kunna DNA polymerase.A lokaci guda, reverse transcriptase baya kunnawa.

PCR ya ƙunshi jerin zagayowar zafi.Kowane zagayowar ya ƙunshi denaturation, annealing da tsawo matakai.

Matakin cirewa ya ƙunshi dumama ɗakin amsawa zuwa digiri 95 ma'aunin celcius da yin amfani da shi don ƙirƙira samfurin DNA mai ɗaure biyu.

A mataki na gaba, ana rage zafin amsawa zuwa digiri 58 a ma'aunin celcius, yana ba da damar na'urar gaba ta share zuwa sashin da ya dace na samfurin DNA ɗin sa guda ɗaya.Matsakaicin zafin jiki kai tsaye ya dogara da tsayi da abun da ke ciki na firamare.

A cikin matakin tsawaita, DNA polymerase yana haɗa sabon madaidaicin DNA wanda ya dace da madaidaicin samfurin DNA.Ta ƙara free nuclei complementary zuwa samfuri a cikin 5'to 3'direction daga dauki cakuda.Zazzabi na wannan mataki ya dogara da DNA polymerase da aka yi amfani da shi.

Bayan zagayowar farko, ana samun maƙasudin DNA mai madauri biyu.

Sa'an nan, shigar da sake zagayowar na biyu.An hana DNA mai madauri biyu don samar da kwayoyin halittar DNA guda biyu.

A mataki na gaba, ana rage zafin da ake yi, ana share abubuwan farko zuwa kowane samfuri na DNA mai igiya guda ɗaya, kuma an goge binciken Taq-man zuwa sashin da aka yi niyya na DNA.

Binciken TaqMan ya ƙunshi nau'in fluorophore wanda aka haɗa shi da 5'ƙarshen binciken oligonucleotide.Lokacin farin ciki da tushen hasken mai hawan keke, fluorophore yana fitar da haske.Bugu da kari, binciken yana kunshe da quencher a 3′ karshen.Matsakaicin jinsin mai ba da rahoto zuwa mai kashewa yana hana gano haske.

A cikin matakin tsawaita, DNA polymerase yana haɗa sabon madauri.Lokacin da polymerase ya isa binciken TaqMan, ayyukansa na 5′nuclease na ƙarshe ya raba binciken, yana raba rini daga mai kashewa.

Tare da kowane sake zagayowar PCR, ana fitar da ƙarin ƙwayoyin rini, wanda ke haifar da haɓakar ƙarfin walƙiya daidai da adadin amplicons da aka haɗa.

Wannan hanya tana ba da damar ƙididdige adadin jerin da aka bayar a cikin samfurin.Adadin gutsuwar DNA guda biyu yana ninka sau biyu a kowane zagayowar.Saboda haka, ana iya amfani da PCR don nazarin ƙananan samfurori.

Don auna sigina mai kyalli, fitilar halogen tungsten, matattarar tashin hankali, mai haskakawa, ruwan tabarau, tacewa da caji tare da na'urar amfani da kyamarar CCD.

MATAKI 4 Gane

Don auna sigina mai kyalli, fitilar halogen tungsten, matattarar tashin hankali, mai haskakawa, ruwan tabarau, tacewa da caji tare da na'urar amfani da kyamarar CCD.

Hasken da aka tace daga fitilar yana nunawa ta hanyar mai haskakawa, yana wucewa ta cikin ruwan tabarau na condenser, kuma yana mai da hankali ga tsakiyar kowane rami.Sa'an nan kuma hasken da ke fitowa daga ramin yana fitowa daga madubi, ya wuce ta hanyar tacewa, kuma kyamarar CCD ta gano.A cikin kowane zagayowar PCR, CCD na iya gano hasken fluorophore mai jin daɗin kai.

Yana canza hasken da aka kama zuwa bayanan dijital.Ana kiran wannan hanyar PCR na ainihin lokaci, kuma tana ba da damar saka idanu na ainihin lokacin ci gaban halayen PCR.