Immobilization of DNA in polyacrylamide gel for the manufacture of DNA and DNA-oligonucleotide microchips.

نویسندگان

  • D Proudnikov
  • E Timofeev
  • A Mirzabekov
چکیده

Activated DNA was immobilized in aldehyde-containing polyacrylamide gel for use in manufacturing the MAGIChip (microarrays of gel-immobilized compounds on a chip). First, abasic sites were generated in DNA by partial acidic depurination. Amino groups were then introduced into the abasic sites by reaction with ethylenediamine and reduction of the aldimine bonds formed. It was found that DNA could be fragmented at the site of amino group incorporation or preserved mostly unfragmented. In similar reactions, both amino-DNA and amino-oligonucleotides were attached through their amines to polyacrylamide gel derivatized with aldehyde groups. Single- and double-stranded DNA of 40 to 972 nucleotides or base pairs were immobilized on the gel pads to manufacture a DNA microchip. The microchip was hybridized with fluorescently labeled DNA-specific oligonucleotide probes. This procedure for immobilization of amino compounds was used to manufacture MAGIChips containing both DNA and oligonucleotides.

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Manual manufacturing of oligonucleotide, DNA, and protein microchips.

A simple procedure for manufacturing microchips containing various gel-immobilized compounds is described. A gel photopolymerization technique is introduced to produce micromatrices of polyacrylamide gel pads (25 x 25 x 20 microm and larger) separated by a hydrophobic glass surface. A pin device for the manual application of a compound in solution onto the activated polyacrylamide gel pad for i...

متن کامل

Fractionation, phosphorylation and ligation on oligonucleotide microchips to enhance sequencing by hybridization.

Oligonucleotide microchips are manufactured by immobilizing presynthesized oligonucleotides within 0.1 x 0.1 x 0.02 mm or 1 x 1 x 0.02 mm polyacrylamide gel pads arranged on the surface of a microscope slide. The gel pads are separated from each other by hydrophobic glass spacers and serve as a kind of 'microtest tube' of 200 pl or 20 nl volume, respectively. Fractionation of single-stranded DN...

متن کامل

High performance liquid chromatographic analysis of reduction products of a thiolated DNA for immobilization on gold nanoparticles

DNA-based nano-biosensors are emerging scope in the field of biosensors. Many synthetic single stranded functional DNAs have been applied for development of such sensors, recently. Immobilization of DNA oligonucleotides on the surface of gold nanoparticles is a key step in the development of most colorimetric nano-biosensors. The bound DNA is usually thiolated and forms Au-S covalent bond to th...

متن کامل

Direct DNA Immobilization onto a Carbon Nanotube Modified Electrode: Study on the Influence of pH and Ionic Strength

Over the past years, DNA biosensors have been developed to analyze DNA interaction and damage that have important applications in biotechnological researches. The immobilization of DNA onto a substrate is one key step for construction of DNA electrochemical biosensors. In this report, a direct approach has been described for immobilization of single strand DNA onto carboxylic acid-functionalize...

متن کامل

Construction of a Synthetic Vector for Preparation of a 100 Base Pair DNA Ladder

DNA size markers are widely used to estimate the size of DNA samples on agarose or polyacrylamide gelelectrophoresis (PAGE). DNA markers can be prepared by mixing PCR products with definite sizes.Alternatively, they are prepared by restriction enzyme digestion of the genomic DNA of bacteriophages ornatural and synthetic DNA plasmids. The present study describes engineering of ...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

عنوان ژورنال:
  • Analytical biochemistry

دوره 259 1  شماره 

صفحات  -

تاریخ انتشار 1998