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Group of oligonucleotide aptamers for specifically identifying aflatoxin B2

A kind of aflatoxin and oligonucleotide technology, which is applied in the direction of recombinant DNA technology, microbial measurement/inspection, DNA/RNA fragments, etc., to achieve the effects of low preparation cost, stable single-stranded oligonucleotide, and simple operation

Active Publication Date: 2013-03-06
JIANGNAN UNIV
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Problems solved by technology

[0005] In view of the advantages of SELEX technology over traditional antigen-antibody technology, it has higher selectivity, specificity and affinity, and the characteristics of being able to distinguish substances with very similar structures. At present, SELEX technology is mainly used in macromolecules such as whole bacteria and cells. In the screening of small molecules, especially in the application of mycotoxins, there are few mycotoxin aptamers. It is reported that the only aptamers of mycotoxins screened are ochratoxin and fumonisin aptamers. If aflatoxin B can be screened 2 The aptamers will be of great significance to the detection of mycotoxins, and will also promote the development of SELEX technology in the field of small molecules, which is also the improvement of SELEX technology itself.

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  • Group of oligonucleotide aptamers for specifically identifying aflatoxin B2
  • Group of oligonucleotide aptamers for specifically identifying aflatoxin B2
  • Group of oligonucleotide aptamers for specifically identifying aflatoxin B2

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Embodiment Construction

[0016] The following is the specific binding part of the aptamer and AFB screened by SELEX technology 2 Dissociation constant nucleic acid aptamer preparation method and rapid detection of some aptamers and AFB 2 applications of dissociation constants.

[0017] 1. Synthesize random single-stranded DNA library and primers (synthesized by IDT)

[0018] Random single-stranded DNA (ssDNA) library: 5'-AGC AGC ACA GAG GTC AGA TG(N40)CCT ATGCGT GCT ACC GTG AA-3', a random ssDNA library with a length of 80 nt was constructed, with fixed primer sequences at both ends and middle It is a random sequence of 40 bases, and the library capacity is more than 1014; Primer I: 5'-AGCAGCACAGAG GTCAGATG-3'; Primer II: 5'-PO 4 -TTC ACG GTA GCA CGC ATA GG-3', the random ssDNA library and the two primers were prepared with TE buffer to make a 100 μmol / L stock solution and stored at -20°C for later use.

[0019] 2. PCR amplification of double-stranded DNA (dsDNA) and single-stranded DNA (ssDNA) lib...

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Abstract

The invention discloses a group of oligonucleotide aptamers for specifically identifying aflatoxin B2. Single-chain DNA (deoxyribonucleic acid) aptamers capable of specifically identifying aflatoxin B2 are obtained through an SELEX (systematic evolution of ligands by exponential enrichment) technology, and can be transferred to report aptamers through a fluorescein label and the like so as to be used for detect aflatoxin B2, and the aptamer sequences have wide application prospect in aspect of accurately and rapidly detecting aflatoxin B2 in food.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to the preparation of a group of aflatoxin B and aflatoxin B using SELEX technology (exponentially enriched ligand system evolution technology) in molecular biology technology 2 High specificity and high affinity nucleic acid aptamer, for the nucleic acid aptamer in the detection of aflatoxin B 2 The application in the paper provides scientific basis and theoretical basis. Background technique [0002] Aflatoxins (AF) are mainly secondary metabolites of Aspergillus flavus and Aspergillus parasitica, and their chemical structures are similar, and they are all derivatives of dihydrofuranocoumarins. At present, the structure of aflatoxin has been determined to be AFB 1 、AFB 2 、AFM 1 and other 18 species, all of which contain difuran ring and oxinone (also known as coumarin) in their basic structures. The former is its toxic structure, and the latter may be related to its carcinogenicit...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/115C12Q1/68
Inventor 王周平王文凤段诺吴世嘉夏雨马小媛
Owner JIANGNAN UNIV
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