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Affinity chromatography of DNA on nonporous copolymerized particles of styrene and glycidyl methacrylate with immobilized polynucleotide
- Source :
- Analytical biochemistry. 312(2)
- Publication Year :
- 2003
-
Abstract
- Nonporous particles of microsize were prepared by the dispersion polymerization of styrene and glycidyl methacrylate and chemically modified to introduce amino groups on the surface by grafting with either hexamethylenediamine or N-methyl-1,3-propanediamine. Aminated particles were then coupled with phosphorylated single-stranded polynucleotides at the 5′-end through covalent linkages. The affinity columns packed with these prepared polynucleotide-immobilized particles effectively retained single-stranded DNA, which could base-pair with the immobilized sequence. Bound DNAs could be eluted to yield a sharp peak by using an aqueous solution of 0.4 M NaOH. The nonspecific adsorption due to the electrostatic interaction between the polynucleotide and the residual amino groups on the particle surface via the amination with hexamethylenediamine was significant and could only be reduced by using a high salt (NaCl) concentration. A higher salt concentration in the elution solution could result in a portion of complementary polynucleotide eluted in the nonretained fraction. However, the nonspecific adsorption of polynucleotides was insignificant in the column packed with DNA-immobilized particles prepared via amination using N-methyl-1,3-propanediamine. The column was effective for microanalysis of sequence-specific DNA.
- Subjects :
- Glycidyl methacrylate
Polymers
Molecular Sequence Data
Polynucleotides
Biophysics
DNA, Single-Stranded
Sodium Chloride
Biochemistry
Chromatography, Affinity
Styrene
Substrate Specificity
chemistry.chemical_compound
Polymer chemistry
Sodium Hydroxide
Molecular Biology
Amination
Dispersion polymerization
Aqueous solution
Base Sequence
Chemistry
Elution
Cell Biology
DNA
Polynucleotide
Hexamethylenediamine
Epoxy Compounds
Methacrylates
Porosity
Subjects
Details
- ISSN :
- 00032697
- Volume :
- 312
- Issue :
- 2
- Database :
- OpenAIRE
- Journal :
- Analytical biochemistry
- Accession number :
- edsair.doi.dedup.....f14db6fa4cf22e9439853dffd13c8731