Specific primer pair for identifying tangerine fruit flies and citrus fruit flies and application thereof

A technique for the identification of B. tangerines and B. tangerines, which is applied in the field of identifying specific primer pairs of B. tangerines and D. tangerines, and can solve problems such as difficulties in morphological identification of B. tangerines.

Active Publication Date: 2019-05-17
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The traditional identification of fruit flies mainly relies on morphological characteristics, and the external morphological characteristics of the two species of Bactrocera are very similar, which can only be determined by the mane sequence of the adult mesothorax (Bactrocera dorsalis has a pair of shoulder plate bristles, and Absence of bristles; Bactrocera citrus has 2 pairs of scapular bristles and 1-2 pairs of upper bristles on forewings) and female ovipositor characteristic

Method used

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  • Specific primer pair for identifying tangerine fruit flies and citrus fruit flies and application thereof
  • Specific primer pair for identifying tangerine fruit flies and citrus fruit flies and application thereof
  • Specific primer pair for identifying tangerine fruit flies and citrus fruit flies and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0108] Embodiment 1, identification of the acquisition of the specific primer pair of Bactrocera citrus and Bactrocera citrus

[0109] 1. Acquisition of the fruit fly mtDNA COI gene sequence

[0110] 1. Test fruit fly samples

[0111] The tested fruit fly samples are shown in Table 1. The samples were all placed in absolute ethanol and stored at -20°C for later use.

[0112] Table 1. Types of fruit fly samples tested and information on collection places

[0113]

[0114]

[0115] 2. PCR amplification

[0116] Using the "Blood / Cell / Tissue Genomic DNA Extraction Kit" (Tiangen Biochemical Technology (Beijing) Co., Ltd. DP304) to extract the genomic DNA of each fruit fly sample in Table 1, refer to the instructions for the specific operation process, and obtain the samples in Table 1 respectively. Genomic DNA of individual fruit fly samples.

[0117] The genomic DNA of each fruit fly sample in Table 1 was used as a template, and the universal primer LCO1490 / HCO2198 was use...

Embodiment 2

[0139] Embodiment 2, identify the specificity detection of the specific primer pair of Bactrocera citrus and Bactrocera citrus

[0140] (1) The inventive method

[0141] One, the specific detection method of the specific primer pair of identifying Bactrocera citrus and Bactrocera citrus

[0142] 1. Test fruit fly samples

[0143] The tested fruit fly samples are shown in Table 4, and all samples were placed in absolute ethanol and stored at -20°C for later use.

[0144] Table 4. Fruit fly species corresponding to conventional PCR reactions

[0145]

[0146]

[0147] 2. PCR amplification

[0148] Using "Blood / Cell / Tissue Genomic DNA Extraction Kit" (Tiangen Biochemical Technology (Beijing) Co., Ltd. DP304) to extract the genomic DNA of each fruit fly sample in Table 4 respectively, refer to the instructions for the specific operation process, and obtain the samples in Table 4 respectively. Genomic DNA of individual fruit fly samples.

[0149] Using the genomic DNA of...

Embodiment 3

[0164] Embodiment 3, identify the sensitivity detection of the specific primer pair of Bactrocera citrus and Bactrocera citrus

[0165] 1. Sensitivity detection method for identifying specific primer pairs of Bactrocera citrus and Bactrocera citrus

[0166] Among the DNA samples of B. dorsalis, one representative population DNA sample from each province and municipality was selected, and one representative population DNA sample from each province and municipality was also selected among the DNA samples of Bacteralis citrus. A total of 6 DNA samples of B. dorsalis were selected from Shaanxi, Hubei, Hunan, Guizhou, Chongqing, and Yunnan; 5 DNA samples of B. dorsalis were selected from Sichuan, Yunnan, Guizhou, Hunan, and Guangxi. The specific information is shown in Table 5.

[0167] Table 5. Information on the origin of Bactrocera citrus and Bactrocera citrus samples

[0168]

[0169] The selected DNA samples were serially diluted to obtain DNA samples with concentrations ...

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Abstract

The invention discloses a specific primer pair for identifying tangerine fruit flies and citrus fruit flies and an application thereof. The invention designs a specific primer pair for identifying thetangerine fruit flies and the citrus fruit flies, and develops a molecular identification technology based on conventional PCR. Experiments prove that the molecular identification technology based onthe conventional PCR can achieve the species identification of the tangerine fruit flies and the citrus fruit flies, and has the advantages of strong specificity, high sensitivity, low cost, simple operation and short time consumption. The primer pair not only solves the problem of identifying the species of non-adult fruit flies and residues of the adult fruit flies, but also provides a practical technology and a theoretical basis for monitoring the epidemic situation of the tangerine fruit flies and citrus fruit flies, distributing and invading of the tangerine fruit flies and the citrus fruit flies in main production areas of China, and subsequent plant quarantine and field management. The primer pair is of great significance to effectively prevent further invasion and diffusion of thetangerine fruit flies and citrus fruit flies, protect agricultural production and ecological security, and promote the development of economic and trade in China.

Description

technical field [0001] The invention belongs to the field of biological technology, and in particular relates to a pair of specific primers for identifying Bactrocera citrus and Bactrocera citrus and application thereof. Background technique [0002] Tephritids belong to Diptera Tephritidae and are widely distributed all over the world. There are about 500 genera and 4,500 species, of which 26 genera and 223 species are economically significant. Fruit flies have a wide range of hosts. While doing damage to economic crops such as fruits and vegetables, they also seriously affect the foreign trade of fruits and vegetables, causing trade barriers and causing huge economic losses. Bactroceratsuneonis and B.minax are two species of oligophagous fruit flies of the subgenus Tetradacus of the genus Bactrocera Bactrocera, which are only harmful to sweet oranges, limes, ponkans, and Wenzhou honey. Citrus crops such as orange, lemon, citron, bergamot, kumquat of the genus Fortunella, ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6888C12N15/11
Inventor 李志红张岳郑林宇
Owner CHINA AGRI UNIV
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