Qhov Tseem Ceeb Qhov txawv - PCR vs DNA Sequencing
PCR thiab DNA sequencing yog ob txoj hauv kev tseem ceeb hauv Molecular Biology. Polymerase Chain Reaction (PCR) yog cov txheej txheem uas tsim ntau cov ntawv luam ntawm DNA tawg. DNA sequencing yog cov txheej txheem uas ua rau kev txiav txim meej ntawm nucleotides ntawm ib qho DNA tawg. Qhov no yog qhov sib txawv tseem ceeb ntawm PCR thiab DNA sequencing. PCR yog ib kauj ruam tseem ceeb koom nrog hauv DNA sequencing.
YPCR yog dab tsi?
Polymerase Chain Reaction (PCR) yog cov txheej txheem DNA amplification siv hauv Molecular Biology. Nws tsim ntau txhiab mus rau tsheej lab ntawm cov ntawv luam ntawm ib qho DNA feem. Txoj kev no tau tsim los ntawm Kary Mullis hauv xyoo 1983. Hauv cov txheej txheem no, cov khoom tawg ntawm DNA yuav tsum tau ua kom nrov nrov ua cov qauv thiab DNA polymerase enzyme ntxiv cov nucleotides ntxiv rau cov primer uas muaj nyob rau hauv PCR sib tov. Thaum kawg ntawm PCR cov tshuaj tiv thaiv, ntau cov ntawv luam ntawm cov qauv DNA yog synthesised.
Muaj ntau yam sib txawv ntawm PCR sib xyaw, suav nrog DNA, DNA polymerase (Taq polymerase), primers (forward thiab thim rov qab primers), nucleotides (lub tsev blocks ntawm DNA) thiab tsis muaj. PCR tshwm sim hauv lub tshuab PCR, thiab cov PCR sib xyaw kom raug yuav tsum tau ntim rau hauv lub tshuab, thiab cov kev pabcuam raug yuav tsum tau tsav. Cov txheej txheem no tso cai rau kev tsim tawm ntau txhiab mus rau ntau lab luam ntawm ib ntu ntawm DNA los ntawm ib qho me me ntawm DNA.
PCR cov tshuaj tiv thaiv tshwm sim nyob rau hauv lub voj voog los tsim cov khoom pom ntawm PCR ntawm cov gel. Muaj peb kauj ruam tseem ceeb koom nrog hauv PCR cov tshuaj tiv thaiv uas yog denaturation, primer annealing thiab strand extension raws li qhia hauv daim duab 01. Peb cov kauj ruam no tshwm sim ntawm peb qhov kub sib txawv. DNA muaj nyob rau hauv ob daim ntawv stranded los ntawm hydrogen bonds ntawm lub complementary bases. Ua ntej yuav cuam tshuam, ob txoj hlua DNA yuav tsum tau muab cais tawm ntawm ib leeg. Nws yog ua los ntawm kev muab qhov kub thiab txias. Thaum qhov kub thiab txias, muab ob npaug rau DNA denature rau hauv ib leeg. Tom qab ntawd cov primers yuav tsum tau los ze zog mus rau lub flanking kawg ntawm lub meej fragment los yog cov noob ntawm DNA. Primer yog ib qho luv luv ntawm ib leeg-stranded DNA uas yog ntxiv rau lub hom phiaj sib lawv liag. Forward thiab thim rov qab primers anneal nrog cov hauv paus ntxiv ntawm qhov kawg ntawm qhov kawg ntawm cov qauv DNA ntawm qhov kub thiab txias. Primers yuav tsum tau tshav kub-resistant. Thaum primers anneal nrog cov qauv DNA, taq polymerase enzyme pib lub synthesis ntawm cov strands tshiab los ntawm kev ntxiv nucleotides uas yog complementary rau lub hom phiaj DNA. Taq polymerase yog lub tshav kub ruaj khov enzyme cais los ntawm cov kab mob thermophilic hu ua Thermus aquaticus. PCR tsis tswj qhov zoo tshaj plaws rau kev ua taq polymerase. Peb theem ntawm PCR cov tshuaj tiv thaiv no rov ua dua los tsim cov khoom PCR uas xav tau. Tom qab txhua qhov tshuaj tiv thaiv PCR, tus lej ntawm DNA daim ntawv yog ob npaug. Yog li ntawd, qhov kev nthuav dav nthuav dav tuaj yeem pom hauv PCR. Cov khoom PCR tuaj yeem pom siv gel electrophoresis thiab tuaj yeem ua kom huv si rau kev kawm ntxiv.
Daim duab 01: Cov kauj ruam tseem ceeb ntawm PCR Reaction
PCR yog cov cuab yeej tseem ceeb hauv kev tshawb fawb txog kev kho mob thiab tshuaj lom neeg. PCR muaj tus nqi tshwj xeeb hauv kev tshawb fawb forensic vim nws tuaj yeem nthuav dav DNA rau cov kev tshawb fawb los ntawm cov qauv me me los ntawm cov neeg ua phem thiab ua cov ntaub ntawv pov thawj DNA. PCR yog dav siv nyob rau hauv ntau qhov chaw ntawm molecular biology suav nrog, genotyping, noob cloning, kev hloov pauv, DNA sequencing, DNA microarrays thiab kuaj txiv neej, thiab lwm yam.
Daim duab 02: Polymerase Chain Reaction
DNA Sequencing yog dab tsi?
DNA sequencing yog qhov kev txiav txim ntawm qhov tseeb ntawm nucleotides - adenine, guanine, cytosine thiab thymine nyob rau hauv ib qho DNA tawg. Cov ntaub ntawv caj ces yog khaws cia rau hauv DNA ua ntu zus los ntawm kev txiav txim kom raug ntawm nucleotides. Yog li, nrhiav qhov tseeb ntawm cov nucleotides hauv DNA tawg yog qhov tseem ceeb heev kom paub txog cov qauv thiab kev ua haujlwm ntawm cov noob.
DNA sequencing raws tu qauv muaj cov txheej txheem sib txawv. Thawj kauj ruam yog kev sib cais ntawm cov neeg nyiam DNA lossis genomic DNA ntawm cov kab mob. Siv PCR (raws li tau piav qhia saum toj no), thaj tsam xav tau ntawm DNA yuav tsum tau nthuav dav. Amplified PCR khoom yuav tsum tau muab cais los ntawm gel electrophoresis thiab purified. Amplified fragments tau txais kev pab raws li cov qauv rau kev ua ntu zus. Sequencing tuaj yeem ua tau raws li Sanger sequencing lossis high throughput sequencing method. Sanger sequencing yuav tsum tau capillary electrophoresis ntawm qhov tshwm sim DNA tawg. Kev txiav txim siab txog qhov tseeb nucleotide tuaj yeem ua tiav los ntawm kev nyeem phau ntawv ntawm autoradiographs lossis siv cov khoom siv DNA automated.
Gene sequencing pab txhawb rau tib neeg genome project thiab ua kom yooj yim daim ntawv qhia ntawm tib neeg genome hauv 2003. Hauv forensics, DNA sequencing enabled qhia txog cov tib neeg uas qhia tshwj xeeb DNA sequences thiab txheeb xyuas cov criminals. Hauv cov tshuaj, DNA sequencing tuaj yeem siv los txheeb xyuas cov noob muaj lub luag haujlwm rau cov caj ces thiab lwm yam kab mob, nrhiav cov noob tsis raug thiab hloov lawv nrog cov noob raug. Hauv kev ua liaj ua teb, DNA sequencing cov ntaub ntawv ntawm qee cov kab mob yog siv los tsim cov qoob loo transgenic nrog cov yam ntxwv xav tau kev lag luam.
Daim duab 03: DNA Sequencing
Qhov txawv ntawm PCR thiab DNA Sequencing yog dab tsi?
PCR vs DNA Sequencing |
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| txheej txheem PCR tsim ntau txhiab mus rau ntau lab daim ntawv theej ntawm cov khoom DNA txaus siab. | DNA sequencing yog txheej txheem ntawm kev txiav txim siab qhov kev txiav txim ntawm nucleotides nyob rau hauv ib tug muab DNA fragment. |
| PCR tsim ntau txhiab rau ntau lab luam ntawm ib qho DNA tshwj xeeb | Qhov no ua rau muaj qhov tseeb ntawm cov hauv paus hauv ib qho DNA feem. |
| Kev koom tes ntawm ddNTPs | |
| PCR tsis xav tau ddNTPs. Nws siv dNTPs. | DNA sequencing yuav tsum tau ddNTPs los txiav tawm strand tsim. |
Summary – PCR vs DNA Sequencing
PCR thiab DNA sequencing yog cov cuab yeej tseem ceeb hauv ntau qhov chaw ntawm Molecular Biology. Kev nthuav dav ntawm DNA tawg yog ua los ntawm cov txheej txheem PCR thaum qhov kev txiav txim kom raug ntawm nucleotides ntawm DNA fragment yog txiav txim los ntawm DNA sequencing. Qhov no yog qhov txawv ntawm PCR thiab DNA sequencing.
