Ma genomic a ma virus ambiri adziwika.Nucleic acid probes omwe ndi magawo afupiafupi a DNA opangidwa kuti asakanizidwe ndi ma virus owonjezera a DNA kapena zigawo za RNA.Polymerase chain reaction (PCR) ndi njira yabwino kwambiri yodziwira ma virus.Njira zowunikira zapamwamba zapangidwa posachedwa.
A. Nucleic acid hybridization njira
Nucleic acid hybridization, makamaka kuphatikiza Kumwera blotting (Kummwera) ndi Kumpoto blotting (Kumpoto), ndi mofulumira kukula njira yatsopano mu munda matenda matenda.Zolinga za hybridization assay ndikugwiritsa ntchito zigawo zazifupi za DNA (zotchedwa "probe") zomwe zimapangidwira kuti zisakanizike ndi zigawo za DNA kapena RNA.Potenthetsa kapena mankhwala amchere, DNA kapena RNA yokhala ndi zingwe ziwiri imagawika kukhala chingwe chimodzi ndiyeno imasasunthika pakuthandizira kolimba.Pambuyo pake, kafukufuku amawonjezedwa ndikusakanikirana ndi DNA kapena RNA yomwe mukufuna.Popeza kafukufukuyo amalembedwa ndi isotopu kapena non-radioactive nuclide, chandamale cha DNA kapena RNA imatha kuzindikirika kudzera mu autoradiography kapena biotin-avidin system.Popeza kuti majeremusi ambiri a ma virus adapangidwa ndikutsatizana, amatha kuzindikirika pogwiritsa ntchito njira zotsatizana ndi ma virus monga ma probe pachitsanzocho.Pakali pano, njira zophatikizira zikuphatikizapo: dot blot , in situ hybridization m'maselo , DNA blotting(DNA) (Southern blotting) ndi RNA blotting(RNA) (Northern blot).
B.PCR Technology
M'zaka zaposachedwa, njira zingapo za in vitro nucleic acid amplification zapangidwa kutengera PCR, kuyesa ma virus osamva kapena osakulitsidwa.PCR ndi njira yomwe imatha kuphatikizira DNA motsatana ndi in vitro polymerase reaction.Njira ya PCR imaphatikizapo kuzungulira kwa kutentha kwa masitepe atatu: denaturation , annealing , ndi extension Pa kutentha kwakukulu (93 ℃ ~ 95 ℃), DNA yamitundu iwiri imasiyanitsidwa ndi zingwe ziwiri za DNA;ndiye pa kutentha kochepa (37 ℃ ~ 60 ℃), ma nucleotide opangira ma nucleotide oyambira anneal ku magawo owonjezera a DNA;pomwe pa kutentha koyenera kwa Taq enzyme (72℃), kaphatikizidwe ka maunyolo atsopano a DNA kumayambira pa primer 3'mapeto pogwiritsa ntchito DNA yowonjezera ngati ma template ndi ma nucleotides amodzi ngati zida.Choncho pambuyo pa kuzungulira kulikonse, unyolo umodzi wa DNA ukhoza kukulitsidwa kukhala maunyolo awiri.Kubwereza ndondomekoyi, unyolo uliwonse wa DNA wopangidwa mumzere umodzi ukhoza kugwiritsidwa ntchito ngati template mumzere wotsatira, ndipo chiwerengero cha maunyolo a DNA chimawirikiza kawiri pamtundu uliwonse, zomwe zikutanthauza kuti kupanga PCR kumakulitsidwa mu liwiro la chipika cha 2n.Pambuyo pa 25 mpaka 30 kuzungulira, kupanga kwa PCR kumadziwika kudzera mu electrophoresis, ndipo mankhwala enieni a DNA amatha kuwonedwa pansi pa kuwala kwa UV (254nm).Chifukwa cha kutsimikizika, kukhudzika, komanso kusavuta, PCR idalandiridwa pakuzindikira matenda ambiri a virus monga HCV, HIV, CMV, ndi HPV.Popeza PCR imakhala yovuta kwambiri, imatha kuzindikira kachilombo ka DNA pamlingo wa fg, opaleshoniyo iyenera kuchitidwa mosamala kwambiri kuti asapewe zolakwika.Kuphatikiza apo, zotsatira zabwino pakuyesa kwa nucleic acid sizitanthauza kuti pali kachilombo koyambitsa matenda.
Pogwiritsa ntchito njira zambiri za PCR, njira zatsopano ndi njira zimapangidwira kutengera njira ya PCR pazolinga zosiyanasiyana.Mwachitsanzo, nthawi yeniyeni ya PCR imatha kuzindikira kuchuluka kwa ma virus;in situ PCR imagwiritsidwa ntchito kuzindikira matenda a virus mu minofu kapena ma cell;PCR yokhala ndi zisa imatha kukulitsa kutsimikizika kwa PCR.Pakati pawo, nthawi yeniyeni yochuluka ya PCR yapangidwa mofulumira kwambiri.Njira zambiri zatsopano, monga TaqMan hydrolysis probe, hybridization probe, ndi ma cell beacon probe, zaphatikizidwa kukhala njira yeniyeni yochulukira ya PCR, yomwe imagwiritsidwa ntchito kwambiri pakufufuza zamankhwala.Kuwonjezera pa kuzindikira kuchuluka kwa ma virus m'madzi a m'thupi la odwala, njira imeneyi ingagwiritsidwenso ntchito pozindikira kuti munthu salolera mankhwala.Chifukwa chake, kuchuluka kwa nthawi yeniyeni kwa PCR kumagwiritsidwa ntchito makamaka pakuwunika kwamankhwala komanso kuyang'anira kulolerana kwamankhwala.
C. Kuzindikira kwakukulu kwa ma viral nucleic acids
Kuti akwaniritse zofunikira zowunikira mwachangu matenda opatsirana omwe angotuluka kumene, njira zingapo zodziwira zowoneka bwino, monga tchipisi ta DNA(DNA), zakhazikitsidwa.Kwa tchipisi ta DNA, ma probe apadera amapangidwa ndikumangidwira ku tchipisi tating'ono ta silicon mu kachulukidwe kwambiri kuti apange DNA probe microarray (DNA) yomwe imatha kusakanizidwa ndi zitsanzo.Chizindikiro cha hybridization chikhoza kujambulidwa ndi ma microscope kapena laser scanner ndikusinthidwanso ndi makompyuta ndipo deta yaikulu yokhudzana ndi majini osiyanasiyana ingapezeke.Pali mitundu iwiri ya DNA chip."Chip synthesis" ili motere: oligonucleotides enieni amapangidwa mwachindunji pa tchipisi.Chinanso ndi DNA pool chip.Ma gene kapena zinthu za PCR zimasindikizidwa mwadongosolo pa slide.Ubwino waukadaulo wa DNA chip ndikuzindikira munthawi yomweyo kuchuluka kwamitundu yambiri ya DNA.Chip chaposachedwa kwambiri chozindikira tizilombo toyambitsa matenda chimatha kuzindikira ma virus a anthu opitilira 1700 nthawi imodzi.Ukadaulo wa chip wa DNA wathetsa zovuta zamachitidwe achikhalidwe cha nucleic acid hybridization ndipo umagwira ntchito motakata kwambiri pakuzindikira matenda a virus komanso maphunziro a epidemiological.
Nthawi yotumiza: Dec-23-2020