Tatelano ea genomic ea likokoana-hloko tse ngata e tsejoa.Nucleic acid probes e leng likarolo tse khutšoane tsa DNA tse etselitsoeng ho kopanya ka likarolo tse tlatselletsang tsa DNA kapa RNA.Polymerase chain reaction (PCR) ke mokhoa o sebetsang haholoanyane oa ho lemoha vaerase.Haufinyane tjena ho entsoe mekhoa ea tlhahlobo e phahameng ea tšebetso.
A. Nucleic acid hybridization technique
Nucleic acid hybridization, haholo-holo e kenyeletsang Southern blotting(Southern) le Northern blotting(Leboa), ke mokhoa o mocha o ntseng o tsoela pele ka potlako lefapheng la tlhahlobo ea kokoana-hloko.Sepheo sa tlhahlobo ea hybridization ke ho sebelisa likarolo tse khutšoane tsa DNA (tse bitsoang "probe") tse etselitsoeng ho kopanya ka likarolo tse tlatsetsang tsa DNA kapa RNA.Ka ho futhumatsa kapa ka kalafo ea alkaline, DNA kapa RNA e nang le likhoele tse peli e aroloa ka likhoele tse le 'ngoe ebe e emisoa ka tšehetso e tiileng.Ka mor'a moo, ho kenngoa lipatlisiso le ho kopanngoa ka sepheo sa DNA kapa RNA.Ha probe e ngotsoe ka isotope kapa non-radioactive nuclide, sepheo sa DNA kapa RNA se ka bonoa ka autoradiography kapa biotin-avidin system .Kaha boholo ba liphatsa tsa lefutso tsa kokoana-hloko li entsoe ka mokhoa o tšoanang le oa tatellano, li ka bonoa ho sebelisoa tatelano e khethehileng ea kokoana-hloko e le li-probes setšoantšong.Hajoale, mekhoa ea hybridization e kenyelletsa: blot blot , in situ hybridization in cell , DNA blotting(DNA) (Southern blotting) le RNA blotting(RNA) (Northern blotting).
Theknoloji ea B.PCR
Lilemong tsa morao tjena, letoto la mekhoa ea in vitro nucleic acid amplification e entsoe ho ipapisitse le PCR, ho lekola livaerase tse sa utloeleng kapa tse ke keng tsa lengoa.PCR ke mokhoa o ka kopanyang tatellano e khethehileng ea DNA ka in vitro polymerase reaction.Ts'ebetso ea PCR e kenyelletsa potoloho ea mocheso oa mehato e meraro: denaturation , annealing , and extension Ka mocheso o phahameng (93 ℃ ~ 95 ℃), DNA e habeli e arotsoe ka likhoele tse peli tsa DNA;ebe ka mocheso o tlase (37 ℃ ~ 60 ℃), li-primers tse peli tsa synthesized nucleotide anneal ho likarolo tse tlatsanang tsa DNA;athe mocheso o loketseng bakeng sa enzyme ea Taq (72℃), kopanyo ea liketane tse ncha tsa DNA e qala ho tloha ho primer 3'end e sebelisa DNA e tlatselletsang joalo ka litempele le li-nucleotide tse le 'ngoe e le lisebelisoa.Kahoo ka mor'a potoloho e 'ngoe le e' ngoe, ketane e le 'ngoe ea DNA e ka holisoa ka liketane tse peli.Ho pheta ts'ebetso ena, ketane e 'ngoe le e' ngoe ea DNA e entsoeng ka potoloho e le 'ngoe e ka sebelisoa e le template potolohong e latelang,' me palo ea liketane tsa DNA e imena habeli potolohong e 'ngoe le e' ngoe, ho bolelang hore tlhahiso ea PCR e matlafatsoa ka lebelo la 2n log.Kamora 25 ho 30 cycles, tlhahiso ea PCR e tsejoa ka electrophoresis, 'me lihlahisoa tse khethehileng tsa DNA li ka bonoa tlas'a leseli la UV (254nm).Bakeng sa molemo oa eona oa ho hlaka, kutloisiso, le boiketlo, PCR e amohetsoe tlhahlobo ea bongaka ea mafu a mangata a kokoana-hloko a kang HCV, HIV, CMV, le HPV.Kaha PCR e na le kutloelo-bohloko haholo, e khona ho bona DNA ea kokoana-hloko boemong ba fg, ts'ebetso e lokela ho etsoa ka hloko haholo ho qoba phoso e fosahetseng.Ho feta moo, sephetho se nepahetseng sa tlhahlobo ea nucleic acid ha se bolele hore ho na le vaerase e tšoaetsanoang sampoleng.
Ka ts'ebeliso e pharalletseng ea mokhoa oa PCR, mekhoa le mekhoa e mecha e nts'etsopele e ipapisitse le mokhoa oa PCR molemong oa liteko tse fapaneng.Mohlala, PCR ea nako ea 'nete e khona ho bona bongata ba vaerase;in situ PCR e sebelisetsoa ho khetholla tšoaetso ea kokoana-hloko liseleng kapa liseleng;PCR ea sehlaha e ka eketsa boleng ba PCR.Har'a bona, PCR ea nako ea 'nete e ntlafalitsoe ka potlako.Mekhoa e mengata e mecha, joalo ka TaqMan hydrolysis probe, hybridization probe, le molecular beacon probe, e kopantsoe ho ba mokhoa oa 'nete oa nako ea PCR, o sebelisoang haholo lipatlisisong tsa bongaka.Ntle le ho khetholla bongata ba kokoana-hloko maling a ’mele oa bakuli ka nepo, mokhoa ona o ka boela oa sebelisoa ho lemoha phetoho e mamellang lithethefatsi.Ka hona, PCR ea nako ea 'nete e sebelisoa haholo tlhahlobong ea phello ea pholiso le tlhahlobo ea mamello ea lithethefatsi.
C. Ho lemoha ka mokhoa o phahameng oa li-viral nucleic acids
Ho fihlela litlhoko tsa tlhahlobo e potlakileng ea mafu a tšoaetsanoang a macha, ho thehiloe mekhoa e fapaneng ea tlhahlobo e phahameng, joalo ka DNA chips(DNA).Bakeng sa li-chips tsa DNA, li-probes tse khethehileng lia etsoa 'me li hokelloa ho li-silicon chips tse teteaneng haholo ho etsa DNA probe microarray (DNA) e ka kopanngoang ka sampole.Letšoao la hybridization le ka bonoa ka microscope kapa laser scanner mme la sebetsoa hape ke komporo mme ho ka fumanoa data e kholo mabapi le mefuta e fapaneng ea lefutso.Ho na le mefuta e 'meli ea DNA chip."Chip synthesis" ke e latelang: li-oligonucleotide tse khethehileng li entsoe ka kotloloho ho li-chips.E 'ngoe ke DNA pool chip.Liphatsa tsa lefutso kapa lihlahisoa tsa PCR li hatisoa ka tatellano selaeteng.Molemo oa theknoloji ea DNA chip ke ho lemoha ka nako e le 'ngoe palo e kholo ea tatellano ea DNA.Mofuta oa morao-rao oa chip ea ho lemoha pathogen e ka khetholla livaerase tsa batho tse fetang 1700 hang.Theknoloji ea DNA chip e rarollotse mathata a mekhoa ea setso ea nucleic acid hybridization 'me e na le ts'ebeliso e pharaletseng haholo ho hlahloba likokoana-hloko le thuto ea mafu a seoa.
Nako ea poso: Dec-23-2020