DNA bar code standard detection fragment 16S rRNA for identifying American cockroach, kit and use method thereof

A standard detection technology for Periplaneta americana, applied in DNA/RNA fragments, recombinant DNA technology, microbial measurement/inspection, etc., to achieve strong specificity and high sensitivity

Inactive Publication Date: 2018-12-07
DALI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is currently no effective DNA barcode identification method with high accuracy.

Method used

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  • DNA bar code standard detection fragment 16S rRNA for identifying American cockroach, kit and use method thereof
  • DNA bar code standard detection fragment 16S rRNA for identifying American cockroach, kit and use method thereof
  • DNA bar code standard detection fragment 16S rRNA for identifying American cockroach, kit and use method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Access to DNA Barcoding Standard Genes for Periplaneta americana

[0062] 1. Sample acquisition

[0063] The adults of Periplaneta americana raised in the standardized breeding base of the national and local joint engineering research center for the development of special medicinal insects (Yunnan Province) were identified as Periplaneta americana by Professor Yang Zizhong of Dali University according to the morphological method. Linn.). After the identification was correct, part of the muscle tissue was extracted from fresh specimens and stored in absolute ethanol.

[0064] 2. Preparation of experimental instruments and reagents

[0065] 2.1 Experimental instruments and reagents

[0066] HWS12 constant temperature electric heating constant temperature water bath (Shanghai Yiheng Technology Co., Ltd., serial number 131022035);

[0067] SIGMA 1-14ED centrifuge (Sigma, Germany, serial number 133828);

[0068] DYCP-31DN electrophoresis instrument (Beijing Liuyi Instru...

Embodiment 2

[0132] Embodiment 2 is used to distinguish the kit of American cockroach

[0133] Positive control: adults of Periplaneta americana;

[0134] Primer pair: forward primer and reverse primer synthesized in embodiment 1, wherein the concentration of forward primer and reverse primer is 10 μ mol / L;

[0135] PCR reaction mixture: 0.1U Taq polymerase, 500 μmol / L dNTP, 20 mmol / L Tris-HCl with pH=8.3, 100 mmol / L potassium chloride and 3 μmol / L magnesium chloride;

[0136] 38 parts of distilled water.

Embodiment 3

[0138] Identification of samples of different forms and processed products of Periplaneta americana

[0139] 1. Collection and storage of samples to be tested:

[0140] The samples to be tested were Periplaneta americana bred in batches at the medicinal insect standardized breeding base under the National and Local Joint Engineering Research Center for the Development of Special Medicinal Insects in Dali Bai Autonomous Prefecture, Yunnan Province, including:

[0141] The 1-day-old larvae, larvae, and adults that had just come out of their egg sheaths, as well as the processed imagoes that had been quickly dried in an oven, and adults purchased from the medicinal material market and identified as Periplaneta americana by Professor Yang Zizhong of Dali University were positive control samples.

[0142] Periplaneta americana has been reviewed by authoritative experts before mass breeding. Part of the muscle tissue of the positive control sample and the sample to be tested were n...

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Abstract

The invention relates to the field of species identification and provides a kit for identifying American cockroach. The kit comprises a positive control, positive and negative primers, a PCR reactionmixture and distilled water; the positive control adopts a DNA bar code standard detection fragment 16S rRNA as shown in SEQ ID NO.1; the sequences of the positive and negative primers are as shown inSEQ ID NO.2 and SEQ ID NO.3. According to the invention, the American cockroach and other similar species can be accurately identified through primer design and PCR amplification, in the manner of finding the 16S rRNA gene fragment (SEQ ID NO.1) from American cockroach, wherein the homology of the fragment on the American cockroach species is 98-100% but the homology with the genes of non-American cockroach is obviously reduced. The kit provided by the invention can be used for identifying the powdered or processed American cockroach, the detection scope is wide, the detection accuracy is higher and the identification time is short.

Description

technical field [0001] The invention relates to the technical field of species identification, in particular to a kit for identifying Periplaneta americana and a use method thereof. Background technique [0002] The American cockroach (Periplaneta americana Linn.) is a long-term used animal medicine in the folk. It is often used as dried or fresh adults as medicine. In recent years, studies have shown that the clinical application records of Periplaneta americana as an insect drug have shown that it has pharmacological effects such as anti-tumor, treatment of cardiovascular diseases, promotion of tissue repair, antibacterial, anti-virus, anti-oxidation, and immune enhancement. Pharmaceutical preparations of Periplaneta americana It is also more and more widely used clinically, and the authenticity identification of Periplaneta americana medicinal materials is the basis for medicinal use and pharmacological research. [0003] At present, the species identification of insects...

Claims

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Application Information

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Patent Type & AuthorityApplications(China)
IPC IPC(8): C12Q1/6888C12Q1/686C12N15/11
CPCC12Q1/686C12Q1/6888C12Q2600/166C12Q2545/113C12Q2565/125
Inventor张成桂李筱玲邵维在刘光明杜雯雯赵微李辉杜一民
OwnerDALI UNIV