کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
7613237 | 1493587 | 2014 | 7 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Coupling of deoxyribonucleic acid to solid supports using 3â² terminal ribose incorporation
دانلود مقاله + سفارش ترجمه
دانلود مقاله ISI انگلیسی
رایگان برای ایرانیان
کلمات کلیدی
موضوعات مرتبط
مهندسی و علوم پایه
شیمی
شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
To develop a new form of DNA coupling under mild reaction and coupling conditions, DNA oligonucleotides were synthesized containing a 3â² ribonucleotide. Upon reaction with millimolar sodium metaperiodate (NaIO4), the ribose is oxidized to a dialdehyde at pH 6.8. This reaction is complete in 30Â min, is quenched with millimolar sodium metabisulfite (Na2S2O5) and is then suitable for coupling to hydrazide-agarose supports. Coupling occurs with a half-time of 27Â min and 80% couples in 2Â h. The EP18 oligonucleotide which binds to the CAAT enhancer binding protein (C/EBP) was synthesized with a 3â² ribose (rEP18) and coupled to hydrazide-agarose. The columns prepared show no significant loss of the oligonucleotide after 50 days. A crude bacterial extract from cells expressing a chimeric fusion protein of GFP-C/EBP was applied to the columns and eluted with different salt concentrations. The active protein elutes in 0.5Â M NaCl and SDS-PAGE/silver stained gels show a single major band which comigrates with GFP-C/EBP as well as three minor contaminants. This provides a new alternative way of coupling DNA to solid supports using mild chemistry which is non-detrimental to the DNA and can be performed if required in the presence of nuclear extract.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Chromatography A - Volume 1339, 25 April 2014, Pages 73-79
Journal: Journal of Chromatography A - Volume 1339, 25 April 2014, Pages 73-79
نویسندگان
Yinshan Jia, Oleg Larionov, Harry W. Jarrett,