کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
4989784 1456932 2017 26 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Plasmid purification by using a new naphthalene tripodal support
ترجمه فارسی عنوان
پاکسازی پلاسمید با استفاده از یک سه پایه جدید نفتالین
موضوعات مرتبط
مهندسی و علوم پایه مهندسی شیمی تصفیه و جداسازی
چکیده انگلیسی
The naphthalene tripodal ligand was synthesized and characterized to assess its purity and subsequently immobilized onto an epoxy-activated Sepharose CL-6B, using mild conditions and resulting in a ligand density of 0.32 mmol naphthalene tripodal/g derivatized Sepharose CL-6B. The complete characterization of naphthalene tripodal Sepharose CL-6B support was performed by High Resolution Magic Angle Spinning (HR-MAS) NMR spectroscopy, scanning electron microscopy (SEM) and elemental analysis. The affinity was measured by SPR biosensor between naphthalene tripodal ligand immobilized on the surface and sc pVAX1-LacZ and the KD was 8.65 × 10−8 ± 1.0 × 10−8 M in 10 mM Tris-HCl pH 8.0, at T = 25 °C, indicating a high affinity. For comparison reasons, the affinity ligand 3,8-diamino-6-phenylphenanthridine (DAPP) was also immobilized on the chip surface and the KD for sc pVAX1-LacZ is lower than with naphthalene tripodal. Saturation transfer difference-nuclear magnetic resonance spectroscopy (STD-NMR) experiments showed that the interactions between the naphthalene tripodal-Sepharose CL-6B and DAPP-Sepharose supports and the 5′-mononucleotides are mainly hydrophobic and π-π stacking. The isolation of sc pDNA isoform was achieved with low salt concentrations, using 95 mM NaCl in binding step and 550 mM NaCl in elution step at T = 4 °C and pH 8, thus reducing the economic and environmental impact.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Separation and Purification Technology - Volume 188, 29 November 2017, Pages 81-89
نویسندگان
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