Oligonucleotide aptamer for specifically identifying patulin

A technology of patulin and oligonucleotides, which is applied in the preparation of test samples, DNA/RNA fragments, material inspection products, etc., can solve the problems of high detection cost, complicated operation, low detection limit, etc., and achieve Stable chemical properties, good reproducibility, and convenient results

A technology of patulin and oligonucleotides, which is applied in the preparation of test samples, DNA/RNA fragments, material inspection products, etc., can solve the problems of high detection cost, complicated operation, low detection limit, etc., and achieve Stable chemical properties, good reproducibility, and convenient results

CN104593374AActive Publication Date: 2015-05-06JIANGNAN UNIV
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  • Oligonucleotide aptamer for specifically identifying patulin
  • Oligonucleotide aptamer for specifically identifying patulin
  • Oligonucleotide aptamer for specifically identifying patulin

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0012] The following is a further description of screening nucleic acid aptamers that specifically bind to patulin by SELEX technology.

[0013] 1. Synthesize random single-stranded DNA library and primers (synthesized by IDT)

[0014] Random single-stranded DNA (ssDNA) library: 5′-CAGCTCAGAAGCTTGATCCT(N40)GACTCGAAGTCGTGCATCTG-3′, a random ssDNA library with a length of 80 nt was constructed, with fixed primer sequences at both ends and a random sequence of 40 bases in the middle, the library capacity for 10 14 Above; upstream primer Ⅰ: 5′-CAGCTCAGAAGCTTGATCCT-3′; downstream primer Ⅱ: 5′-CAGATGCACGACTTCGAGTC-3′, the random ssDNA library and the two primers were prepared into 100 μmol / L stock solution with TE buffer and stored at -20°C spare.

[0015] 2. Preparation and purification of ssDNA

[0016] The reaction system is: template DNA 5 μL, FAM-labeled upstream primer 1 μL (20 mM), phosphorylated downstream primer 0.5 μL (20 mM), dNTPmix (25 mM) 0.5 μL, 10×PCR amplificatio...

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Abstract

The invention discloses an oligonucleotide aptamer for specifically identifying patulin. Through a graphene oxide-based systematic evolution technology of ligands by exponential enrichment, a single stranded oligonucleotide aptamer capable of specifically identifying patulin is acquired. The aptamer can be converted into a report aptamer by marking functional groups and the like. The aptamer is used for detecting patulin. The sequence of the aptamer has a wide application prospect in accurately and quickly detecting patulin in a food.

Description

technical field [0001] The present invention relates to the field of biotechnology, in particular to the preparation of a nucleic acid aptamer with high specificity and high affinity to patulin by using a SELEX technology (exponentially enriched ligand system evolution technology). The ligand provides the basis for the rapid detection of patulin. Background technique [0002] Patulin is a fungal metabolite produced by the genera Penicillium, Aspergillus, and Clothes, and is mainly found in apples and their products. Patulin is stable to heat, its structure cannot be destroyed by heating at 100°C for 15 minutes, and it can be preserved for a long time in acidic solution and organic solvent. Patulin has been used as an antibiotic because it inhibits the growth of a variety of bacteria. It has acute toxicity such as convulsions, intestinal bleeding, pulmonary hemorrhage, and kidney damage. It also has neurotoxicity, immunotoxicity, genotoxicity, and teratogenicity. , carcinog...

Claims

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Application Information

Patent Timeline
06 May 2015
Publication
CN104593374A
IPC
C12N15/115; G01N33/53; G01N1/40
Inventors
王周平; 张维潇