Hippodamia variegate specificity COI primer, kit containing primer and detection method thereof

A technology of lady beetle polymorpha and a detection method, applied in the field of molecular biology, can solve the problems of complex biological control effect of population dynamic pests, influence, inability to accurately assess the predation effect of lady beetles and lady beetles, etc., and achieves high practical value and simple operation. , the effect of high sensitivity

Inactive Publication Date: 2015-07-22
INST OF PLANT PROTECTION CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

With the in-depth study of this effect, researchers found that it has a complex and huge impact on the population dynamics of different species in the farmland ecosystem and on the effect of biological control of pests
In terms of research methodology, it is found that traditional research methods such as routine intestinal anatomy and behavioral observation cannot accurately assess the predation among ladybugs in the ladybug group
At present, no specific detection method for polydactyly has yet to be established. In the field of species identification, the polymerase chain reaction (PCR) technology occupies an important position because of its strong specificity, high sensitivity, simplicity and rapidity. important position

Method used

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  • Hippodamia variegate specificity COI primer, kit containing primer and detection method thereof
  • Hippodamia variegate specificity COI primer, kit containing primer and detection method thereof
  • Hippodamia variegate specificity COI primer, kit containing primer and detection method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Amplification Effect of Primer HvF1 / HvR1 on Ladybug polydactylus

[0024] 1. Preparation of the polydactylyx genome

[0025] Put a single ladybug in a 1.5mL centrifuge tube to grind it, and use TIANamp Genomic DNA Kit blood / cell / tissue genomic DNA extraction kit (spin column type) (Tiangen Biological Co., Ltd., Beijing) to extract single The head ladybug genome. The DNA solution was stored at -20°C for later use, and 1 μL of the solution was taken as a DNA template during PCR amplification.

[0026] 2. Synthesis of specific COI primers for the detection of polyisomysis

[0027] The specific COI primer sequences are as follows:

[0028] HvF1: 5'-GGAAGATTAATTGGAAATGAT-3' (SEQ ID No.1)

[0029] HvR1: 5'-CCTATTTCAACAATTCTTCTAAAT-3' (SEQ ID No.2)

[0030] 3. PCR amplification

[0031] The reaction system is 20 μL, including: 10×EasyTaq buffer 2.0 μL, dNTP (2.5 mM) 0.4 μL, EasyTaq DNA polymerase (5U / μL) 0.2 μL, forward and reverse primers (10 μM) each 0.4 μL, ...

Embodiment 2

[0040] Example 2 Determination of Primer HvF1 / HvR1 on the Minimum Detection Amount of Ladybug polydactylus

[0041] 1. Preparation of polydactylyx template DNA

[0042] According to Example 1, the genomic DNA of single-headed ladybug polyheterosus was extracted, and then the original template solution (the concentration of the DNA solution was 3ng / μL) was diluted in a descending gradient by 2 times until no band could be detected, and 1 μL was used as a template for PCR amplification. Add directly to the PCR reaction system, and the reaction system is the same as that described in Example 1.

[0043] Utilize the primer HvF1 / HvR1 to do the determination of the minimum detection amount, carry out PCR amplification with different dilution multiples of the polydactyly ladybug genomic DNA as the template, the DNA concentration detection limit is 1.5ng / μL ( figure 2 ).

[0044] 2. Preparation of Plasmid of polyisomysis

[0045] According to Example 1, the genomic DNA of ladybug ...

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Abstract

The invention relates to a hippodamia variegate specificity COI primer, and a detection kit containing the primer. A pair of hippodamia variegate specificity COI primers HvF1 and HvR1 (SEQ ID No.1 and 2) is designed according to a specific mt deoxyribonucleic acid (DNA) sequence of hippodamia variegate. The pair of primers just has specific amplification capability on the hippodamia variegate; the amplification product is 230bp; the plasmid concentration detection limit is 1.503567837E+05; the DNA concentration detection limit is 1.5ng / mu L. By adopting a polymerase chain reaction (PCR) technology, the detection accuracy is improved, the detection time is saved, and the kit product is applicable to basic popularization and application.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a polydactylyx specific COI primer, a kit containing the primer and a detection method thereof. Background technique [0002] Hippodamia variegate (Goeze) belongs to Insecta, Coleoptera, Coccinidae and Coccinidae subfamily. It often preys on aphids such as cotton aphid, wheat aphid, soybean aphid, and corn aphid, so it is more common in crops such as cotton, wheat, cowpea, and corn. The lady beetle originated in the Palearctic region and is a common aphid predator in Europe, and has been documented in South Africa, Kenya, India, China, Canada, the United States, and Chile. In my country's Xinjiang, Shandong and other places, the ladybug is an important predatory natural enemy of the cotton aphid; in South Africa, it is also very significant in controlling the pests of the wheat aphid. It can be seen that the ladybug is an effective biological control species. [0003] At the s...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/11C12Q1/68
Inventor 杨帆王倩陆宴辉徐建祥
Owner INST OF PLANT PROTECTION CHINESE ACAD OF AGRI SCI
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