Izixhobo zokucoca i-Oligo ye-Oligo primer synthesis

Isixhobo sokucoca i-Oligo ye-Oligo primer synthesis,
umatshini wenkxaso we-oligo synthesis,

Inkcazo

Iindlela ezahlukeneyo zokuCoca

1. Gel Electrophoresis Chromatography Ukuhlanjululwa
Sebenzisa denaturing polyacrylamide gel electrophoresis chromatography ukuhlanjululwa.I-arhente ye-denaturing ngokubanzi yi-4M ye-formamide okanye i-7M urea, i-concentration ye-acrylamide iphakathi kwe-5-15%, kwaye inxalenye ye-methacrylamide iphakathi kwe-2-10%.
Emva kwe-electrophoresis, indawo yebhanti ye-nucleic acid kufuneka imiselwe phantsi kokukhanya kwe-ultraviolet, i-gel equkethe i-nucleic acid ekujoliswe kuyo iyanqunyulwa, i-nucleic acid ityunyuzwe kwaye ihlanjululwe, kwaye ke isisombululo se-leaching sigxininiswe, sichithwa. quantified kwaye lyophilized.

2. DMT-On, HPLC Ukucoca
Khetha imo ye-DMT-On ngexesha le-synthesis, imveliso ekrwada yayiyi-centrifuged kwaye igxininiswe kwiqondo lokushisa ukuze kususwe i-ammonia engaphezulu emva kwe-aminolysis.
Ukwahlula kwenziwa ngokusebenzisa ikholomu ye-C18 kunye ne-acetonitrile kunye ne-10% ye-triethylamine-acetic acid (TEAA) njenge-eluent.Emva kokuba i-elution igqityiwe, igxininiswe, kwaye ke iqela le-DMT lisuswa nge-trifluoroacetic acid.Emva kokungathathi hlangothi, ezinye iityuwa kunye neamolekyu ezincinci zisuswa nge-tube yokunqunyulwa, kwaye ekugqibeleni i-desalted.

Le ndlela inokufumana imveliso enobunyulu obuphezulu, kodwa kuyimfuneko ukunikela ingqalelo kwisiganeko sokuhlanjululwa.

3. DMT-Off, HPLC Ukucoca
Khetha i-DMT-Off ngexesha lokudityaniswa, kunye nemveliso ekrwada yayiyi-centrifuged kwaye igxininiswe kwiqondo lobushushu begumbi ukususa i-ammonia engaphezulu emva kwe-ammonolysis.
Ukwahlula kuqhutywe ngokusebenzisa ikholomu ye-C18 kunye ne-acetonitrile kunye ne-10% ye-triethylamine-acetic acid emanzini njenge-eluent.Emva kokuba ukwahlukana kugqityiwe kwaye kulinganiswe, i-aliquots i-lyophilized.

Le ndlela ifuna ukulungiswa ngononophelo kweemeko zokwahlukana, kwaye inokufumana kwakhona iimolekyuli ekujoliswe kuzo ezisulungekileyo.

izixhobo ze-oligo synthesis accerry zomjikelo we-dna rna synthesis, ofanele i-primer eqhelekileyo, kunye nezinye i-primer chain chain primer, yenza i-oligo icoceke kakhulu.
umenzi waseTshayina we-oligo synthesis izixhobo ezinomgangatho olungileyo kunye nenkonzo engcono.