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Primer pair for detection of idgf4 gene expression in fruit fly

A technology of gene expression and primer pairs, which is applied in biochemical equipment and methods, microbial measurement/inspection, DNA/RNA fragments, etc., can solve the problems of fruit rot and fall off, fungal infection, etc., and achieve simple operation, low cost, The effect of protecting agricultural production and ecological security

Active Publication Date: 2021-10-15
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The harm is that adults lay eggs on the inner side of the peel, and the larvae eat the fruit, causing fungal infection and causing the fruit to rot and fall off

Method used

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  • Primer pair for detection of idgf4 gene expression in fruit fly
  • Primer pair for detection of idgf4 gene expression in fruit fly
  • Primer pair for detection of idgf4 gene expression in fruit fly

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1. Preparation of primer pairs for detection of fruit fly CG5210 and Idgf4 gene expression

[0040] This embodiment provides a pair of primers for detecting the expression levels of CG5210 and Idgf4 genes in fruit flies, which are respectively denoted as CG5210-rt and Idgf4-rt. CG5210-rt is composed of two single-stranded DNAs shown in sequence 1 and sequence 2 in the sequence listing, the molar ratio of the two single-stranded DNAs in the primer pair is 1:1, and the two single-stranded DNAs are packaged independently. Idgf4-rt is composed of two single-stranded DNAs shown in sequence 3 and sequence 4 in the sequence listing, the molar ratio of the two single-stranded DNAs in the primer pair is 1:1, and the two single-stranded DNAs are packaged independently.

Embodiment 2

[0041] Example 2, using the primer pair of Example 1 to detect the expression of CG5210 and Idgf4 genes in fruit flies

[0042] Fruit flies to be tested: Guava fruit flies collected from Yunnan and Bactrocera dorsalis collected from Guangzhou, 50 first-instar larvae, 30 second-instar larvae, and third-instar early larvae (5 days after egg hatching) 30 instar larvae), 20 late third-instar larvae (7-day-old larvae after egg hatching), 6 early-stage pupae (1-day-old pupae since pupation), 6 mid-stage pupae (5-day-old pupae since pupation) 6, 6 late stage pupae (9 days old pupae since pupation), 10 immature adults after eclosion, 10 sexually mature adults 10 days old.

[0043] The total RNA of each fruit fly was extracted using the RNAsimple Total RNA Extraction Kit of Tiangen Biochemical Technology (Beijing) Co., Ltd., and then reverse-transcribed to obtain cDNA. Using the cDNA obtained by reverse transcription of Bactrocera dorsalis and Bactrocera guava as templates, the Takara...

Embodiment 3

[0054] The primers of embodiment 3, embodiment 1 are to the optimization of required template amount

[0055] Carry out 10-fold gradient dilution respectively to the cDNA that embodiment 2 obtains, obtain the cDNA dilution liquid of the dilution 10 times, 100 times, 1000 times and 10000 times of the specific development stage of each fruit fly (the content of cDNA is respectively 100 μ g / μl, 10 μ g / μl, μl, 1 μg / μl and 0.1 μg / μl), and then use this as a template to determine the optimal template amount using the reaction system and reaction procedure in Example 2, and use water to set up a negative control.

[0056] The results showed that using 100 μg / μl, i.e. 10-fold diluted cDNA, as a template, both primer pairs could obtain positive amplification curves in B. dorsalis and B. guava, and the amplification curves of internal reference genes and target genes It is a single peak while satisfying the CT value of the internal reference gene at about 10, indicating that the cDNA qu...

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Abstract

The invention discloses a pair of primers for detecting the expression level of the fruit fly Idgf4 gene. The primer pair disclosed in the present invention consists of two single-stranded DNAs shown in sequence 3 and sequence 4 in the sequence listing. Experiments have proved that the primer pair for detecting the expression of the fruit fly Idgf4 gene provided by the present invention can specifically, quickly and accurately quantitatively detect the Idgf4 gene of Bactrocera dorsalis and Bactrocera guava, and can be used for interference of the fruit fly Idgf4 gene Efficiency evaluation. In addition, quantitative detection solves the problem of comparing the expression of the same gene among different fruit flies, and is convenient, fast and low-cost to operate. The present invention can also provide tools and related targets for the research of the Idgf4 gene and the control of fruit flies, which is of great significance to the control of fruit flies, and is useful for effectively controlling the spread of dangerous fruit flies, protecting agricultural production and ecological safety, and promoting my country's economic and trade development provides potential target gene reference.

Description

technical field [0001] The invention relates to a pair of primers for detecting the expression level of the fruit fly Idgf4 gene in the field of biotechnology. Background technique [0002] Bactrocera dorsalis (Hendel) belongs to Diptera, Tephritidae Tephritidae, and Bactrocera genus Bactrocera. It is a polyphagous pest and can eat more than 250 kinds of fruits and vegetables in 46 families such as citrus, guava, and mango. The larvae feed on the inside of fruits and vegetables and the adults lay eggs on the fruit skin to form holes, causing fruit drop or whole fruit to rot. This not only brings serious harm to fruit and vegetable production, but also has a serious impact on import and export trade. Due to its wide host range, high fecundity and strong adaptability, many countries list it as a key quarantine object (Zhang Bin et al., 2008). [0003] Bactrocera dorsalis is native to the tropical regions of Asia and now has a wide distribution, involving tropical, subtropical...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6888C12Q1/686C12N15/11
CPCC12Q1/686C12Q1/6888C12Q2600/158C12Q2561/113C12Q2563/107
Inventor 李志红顾欣悦柳丽君粟耘
Owner CHINA AGRI UNIV
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