Imigaqo yeNucleic Acid Synthesis

I-nucleic acid synthesis yenziwa kusetyenziswa indlela eqinileyo ye-sialic amide triglyceride, apho i-3′ isiphelo se-DNA singenakushukunyiswa kwi-substrate yesigaba esiqinileyo kunye nee-nucleotides zongezwa kwi-3′ ukuya kwi-5' icala de i-DNA fragment efunwayo yenziwe. .Oku kwahlukile kwi-DNA synthesis ngokusetyenziswa kwe-DNA polymerase.

Ukuphela kwe-3 yesiseko sokuqala singenakunyakaziswa kwi-CPG ngexesha lokudibanisa, i-5'-OH yesiseko esilandelayo ikhuselwe nge-di-p-tolyl trityl DMT, iqela le-amino kwisiseko likhuselwe nge-benzoic acid kunye ne-3 -OH emva koko isebenze kunye ne-amino phosphate compound.Isiseko se-1 se-5 5'-OH yesiseko se-1 kunye ne-3'-OH yesiseko esilandelayo senza i-phosphite triglyceride, ethi emva koko i-oxidised nge-iodine kwi-phosphate triglyceride, umkhuseli kwisiseko sesibini i-5'-OH iyasuswa. Ukongeza i-dichloroacetic acid i-DM imijikelezo ngokongezwa kwesiseko esilandelayo, kwaye emva kokudibanisa umkhuseli kwi-5'-OH isuswe nge-asidi ebuthakathaka Iqhekeza linqanyuliwe kwi-resin eqinile kunye ne-concentrated ammonium hydroxide, umkhuseli ususwe kwisiseko. kunye ne-ammonium hydroxide egxininisiweyo phantsi kokufudumala, i-ammonium hydroxide isusiwe, iqhekeza i-vacuum yomisiwe, kwaye i-nucleic acid ihlanjululwa yi-chromatography yamanzi okanye i-PAGE.

Amanyathelo Oligo Synthesis

Ukuvalwa

II-5 ekupheleni kweqela elikhuselayo le-DMT lisuswa ngokongeza isisombululo se-TCA.

Ukuqaliswa

Ukuxuba i-activator kunye ne-oligonucleotide monomer ukwenza i-oligonucleotide monomer esebenzayo ephakathi.

Ukudibanisa

Iqela le-5-terminal hydroxyl liphendula kunye ne-oligonucleotide esebenzayo ephakathi kwi-anti-generative condensation reaction ukuze yenze i-phosphite triglyceride bond engazinzanga.

Capping 

Ukongezwa kwe-agent ye-capping yenza i-acetylation capping reaction kunye namaqela e-hydroxyl angaphezulu angabandakanyekanga kwi-condensation reaction.

I-oxidation 

Ukongezwa kwesisombululo se-iodine esinamanzi siphendula kunye neebhondi zephosphite ezingazinzanga nge-oxidation ukwenza i-stable phosphoric acid triglyceride bonds.

I-Cyanoethyl Cleavage

Imveliso ye-nucleic acid emva kokuhlanganiswa, kunye nokongezwa kwesisombululo se-DEA ukususa iqela le-cyanoethyl kwi-phosphate triglyceride bond.

Icandekile&Khusela

Amaqhekeza e-nucleic acid acandwa kumphathi wesigaba esiqinileyo ngokongeza isisombululo esigxininisiweyo se-ammonium hydroxide kwaye sifudunyezwe ukususa amaqela akhuselayo kwiibhondi ze-hydrogen zeziseko.