Metotia suesuega faapaiolosi Molecular ma lo latou faaaogaina

faapaiolosi Molecular metotia suesuega ei ai se matafaioi taua i le vailaau po nei, faasaienisi faapitoa, ma paiolo. Faafetai i le aitalafu i le suesuega o le DNA ma RNA, e mafai e le tagata e sailiili le genome o le tino fuafuaina e le sui sooupu causative, e iloa le acid nucleic manao i se tuufaatasiga o acids, ma isi

metotia Molecular-moni. O le a ai?

I tua i le 70-80s o le saienitisi o le uluai e decipher le genome tagata. O lenei mea ua tuuina uunaʻi ai i le atinae o inisinia tau kenera ma paiolo molecular. Suesuega o meatotino a DNA ma RNA ua le uiga e faapea e nei mafai ai ona faaaogaina nei acids nucleic mo le faamoemoe o faamaʻi auga, Gene suesuega.

Sauniuniga o le DNA ma RNA

faapaiolosi Molecular metotia diagnostic manaomia e amata mea: E masani ona lenei acid nucleic. E tele ni auala e filifili nei mea mai le sela o meaola soifua. Taitasi ei ai lona tulaga lelei ma tulaga le lelei, ma e tatau ona iloiloina pe a filifilia se auala o le faaesea o acids nucleic i se faiga mama.

1. Sauniuniga o DNA e Marmur. O le auala e aofia ai le faia o le tuufaatasiga o mea ava malosi, precipitates mafua DNA mama. O le downside o lenei metotia o le faaaogaina o mea e matautia, phenol ma chloroform.

2. Faʻaʻese o DNA i le Boom. O le aano autu lena o loo faaaoga iinei - o thiocyanate guanidine (GuSCN). Ua uunaʻia e le Deposition o acid deoxyribonucleic i substrates faapitoa, lea e mafai ona mulimuli ane aoina e ala i se buffer faapitoa. Ae peitai GuSCN - o se inhibitor o le TCP, ma e oo lava i se vaega itiiti o lena e oo atu teuina i totonu o DNA e mafai ona aafia ai le ala o le tali filifili polymerase, lea e taua pe a galulue faatasi ma acids nucleic.

3. Precipitation o le tulaga le mama. O le auala e ese mai tagata muamua i lena le molecules i latou lava e le o teuina i dehoksiribonukleinovoy acid ma tulaga le mama. Ina ia faia lenei mea, faaaoga le exchangers Ion. O le tulaga le lelei o le le mea uma e mafai ona foia.

4. suesuega Misasa. O lenei auala o loo faaaogaina i tulaga e te le manaomia le i ai faamatalaga saʻo e uiga i le fausaga o le molecule DNA, ma le manaomia e aumai se fuainumera. O le mafuaaga e faapea e mafai ona faaleagaina le faatulagaga acid nucleic pe a taulimaina detergents, aemaise alkalies.

Faavasegaina o metotia suesuega

Uma metotia molecular-moni o suesuega o loo vaevaeina i ni vaega tetele e tolu:

1. Amplification (e faaaoga ai se plurality o enzymes). E aofia ai PCR - tali filifili polymerase, lea ei ai se matafaioi taua i le tele o metotia diagnostic.

2. Neamplifikatsionnye. O lenei vaega o auala ua fesootai tonu ma le galuega o le faafefiloi nucleic acid. Faataitaiga e 3 ituaiga blots, i situ hybridization, ma isi

3. Auala e faavae i luga o le le aloaia o le faailoga mai le molecule suʻesuʻega, lea e noatia ai i se suʻesuʻega e DNA po RNA patino. Faataitaiga - hybridization faiga fofo System pueina Hybride (hc2).

Enzymes e mafai ona faaaoga i auala suesuega faapaiolosi molecular

E toatele molecular metotia diagnostic aofia ai le faaaogaina o vaega lautele o enzymes. O loo i lalo o loo faaaogaina le tele o taimi:

1. tapulaa enzyme - "oti" le molecule DNA i le vaega e tatau ai.

2. DNA polymerase - synthesizes a molecule acid deoxyribonucleic lua-paulia.

3. transcriptase faafeagai (suia transcriptase) - faaaogaina e synthesize DNA mai se mamanu RNA.

4. DNA ligase - e nafa ma le faavaeina o phosphodiester sootaga nucleotides.

5. exonuclease - aveesea nucleotides mai le faaiuga vaega o le molecule acid deoxyribonucleic.

PCR - o le auala autu o le amplification DNA

tali filifili Polymerase (PCR) ua lautele faaaogaina i paiolo molecular nei. O lenei auala, lea na a mafai ona maua molecule DNA tasi se aofaiga tele o ni kopi (amplifying molecule).

O galuega tauave autu o PCR:

- auga o faamai;

- cloning vaega DNA, Gene.

E faataunuu ai le tali filifili polymerase manaomia ai le elemene nei: molecule DNA muamua, a polymerase DNA thermostable (Taq po Pfu), deoxyribonucleotide phosphates (punaoa o nofoaga autu o nitrogen), o le primers (2 primer 1 molecule DNA) ma lava se faiga buffer lea e mafai ona faia tali uma.

aofia PCR o laasaga e tolu: denaturation, annealing primer ma elongation.

1. denaturation. I se vevela o 94-95 tikeri Celsius proshodit motusia noataga hydrogen le va o le filifili e lua o le DNA, ma o se taunuuga tatou te maua molecules lua tasi-paulia.

2. primers annealed. I se vevela o 50-60 tikeri centigrade tupu accession primers i le tuluiga o le tasi-paʻulia molecules acid nucleic e tusa ma le ituaiga o complementarity.

3. Elongation. I se vevela o le 72 tikeri ua faaluaina paʻulia molecules acid deoxyribonucleic afafine synthesized.

DNA sequencing

masani ona manaomia ai le malamalama metotia suesuega faapaiolosi Molecular o le faasologa o nucleotides i se molecule o acid deoxyribonucleic. Ina ia iloa le code tuufaasolo faasologa. a faavae diagnostics Molecular o le lumanai i le malamalama e maua i le faaiuga o le faasologa o tagata.

O ituaiga nei o sequencing:

• sequencing e le Maxam-Gilbert;
• sequencing Sanger;
• pyrosequencing;
• sequencing nanoporovoe.