Lub hauv paus ntsiab lus ntawm DNA synthesis

Cov tshuaj DNA synthesis yog ua raws li cov txheej txheem sib xyaw ua ke thiab phosphoramide chemistry.Qhov sib txawv ntawm cov DNA synthesis, cov khoom siv hauv cov tshuaj DNA synthesis yog DMT (4, 4-dimethoxytrityl) thiab phosphoramidite hloov deoxyriboxyside, raws li qhia hauv daim duab 1. Cov DNA synthesis yog nqa tawm ntawm cov khoom txhawb nqa (peb tuaj yeem muab ntau yam ntawmOligo Synthesis Kem), xws li CPG (tswj pore iav) thiab PS (polystyrene), thiab tag nrho cov synthesis yog ua los ntawmDNA/RNA synthesizer, uas yog peb cov khoom siv tseem ceeb, muaj cov qauv sib txawv xws li: HY Ib Leeg Channel Synthesizer, HY-12, HY-192 thiab lwm yam, cov kev taw qhia synthesis yog los ntawm 3' mus rau 5', thiab ib qho nucleotide yog nkag mus rau cov khoom txhawb nqa ntawm ib qho. synthesis voj voog.Lub voj voog kev sib txuas feem ntau muaj plaub kauj ruam, Deprotection, Coupling, Capping thiab Oxidation (Daim duab 2).Lub Deprotection tau teem kom tshem tawm DMT pawg ntawm cov khoom txhawb nqa lossis 5 'hydroxyl pawg ntawm cov nucleoside yav dhau los, 3% trichloroacetic acid hauv dichloromethane yog siv los ua cov tshuaj tiv thaiv kev tiv thaiv.Tom qab ntawd cov phosphoramidite hloov kho deoxyriboxyside raug tso cai rau kis hydroxyl pawg nrog kev pab ntawm activator, piv txwv li 5-ethylthiotetrazole los yog 4, 5-dicyanoimidazole, tsim cov phosphitetriester (III), thiab paub txog cov kauj ruam Coupling.Ib kauj ruam Capping yog ua los thaiv cov pab pawg hydroxyl uas tsis muaj kev cuam tshuam thaum lub sij hawm sib txuas, kom txo qis kev tsim ntawm cov khoom tsis zoo sib xws.Thaum kawg, qhov tsis ruaj khov phosphitetriester (III) yog oxidized rau tshuaj ruaj khov phosphortriester (V) nrog iodine li oxidant nyob rau hauv tam sim no ntawm pyridine.

Lub synthesized DNA yuav cleavage los ntawm cov khoom txhawb nqa los ntawm aminolysis, 2-cyanoethyl tiv thaiv pab pawg ntawm phosphortriester thiab amide ntawm nucleobase yog cleavaged tib lub sijhawm, Synthesis Phaj thiab Synthesis Kem hauv racks raug tso ncaj qha rau hauv cov tshuaj tiv thaiv chamber. ntawmCov cuab yeej tiv thaiv.Crude DNA tuaj yeem npaj los ntawm HPLC, electrophoresis thiab OPC ntawm kev thov, peb kuj tuaj yeem muab rau kojKhoom Siv Purificationrau Post-processing.