Application of migratory locust Rab5 gene and dsRNA thereof in migratory locust prevention and treatment

A technology of migratory locusts and genes, applied in the direction of DNA/RNA fragments, applications, genetic engineering, etc., can solve problems such as death, abnormal development, and abnormal life activities of insects, etc., to achieve strong insecticidal specificity and efficient lethality The effect of high rate, high specificity and lethality

Active Publication Date: 2021-04-16
SHANXI UNIV
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Problems solved by technology

After silencing the target gene, the life activities of insects cannot be carr

Method used

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  • Application of migratory locust Rab5 gene and dsRNA thereof in migratory locust prevention and treatment
  • Application of migratory locust Rab5 gene and dsRNA thereof in migratory locust prevention and treatment

Examples

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Embodiment 1

[0025] Obtaining the Sequence and dsRNA of the Small GTPase Gene 5 (LmRab5) of Migratory Locust

[0026] 1. Acquisition of migratory locust small GTPase gene 5 (LmRab5) sequence

[0027] 1) Search for the sequence of the migratory locust Rab5 gene in the migratory locust transcriptome database

[0028] Based on the migratory locust transcriptome database, bioinformatics methods were used to search for the sequence of the migratory locust Rab5 gene. After sequence analysis and comparison, a total of 1 migratory locust Rab5 gene sequence was obtained.

[0029] 2) Design of primers required for PCR amplification:

[0030] According to the above gene fragment, the upstream primer and downstream primer were designed using primer premier 5.0 software. The sequences of the primers are shown in SEQ ID NO: 3 and SEQ ID NO: 4 respectively. The primers were synthesized by Shanghai Sangon Bioengineering Co., Ltd.

[0031] 3) PCR amplification reaction

[0032] The total RNA of the 3rd ...

Embodiment 2

[0045] The dsRNA Lethal Experiment of Migratory Locust Small GTPase Gene 5 (LmRab5)

[0046] 1. Specific dsRNA Injection

[0047] 2 μl (6 μg) of the dsRNA synthesized in Example 1 above was injected with a 10 μl microsyringe between the second and third abdominal segments of the 2nd-day-old nymphs of the 3rd instar migratory locust, and as the experimental group, a total of 36 were injected, with half male and half male. The control group was injected with the same volume and concentration of dsEGFP. After injection, migratory locusts were raised in a constant temperature culture room at 30°C (light:dark time=14h:10h, temperature 30±2°C, humidity 60%), and the control and treatment groups were fed fresh wheat seedlings and wheat bran every day.

[0048] 2. Silence detection of migratory locust small GTPase gene 5 (LmRab5)

[0049] Collect 6 migratory locusts injected with dsEGFP (control group) and dsLmRab5 (experimental group) for 24 hours each, and extract total RNA from n...

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Abstract

The invention relates to the technical field of biology, in particular to a migratory locust Rab5 gene and application of dsRNA of the migratory locust Rab5 gene in migratory locust prevention and treatment. Specifically, through a bioinformatics method, a fragment of the migratory locust small GTP enzyme gene 5 is obtained from a migratory locust transcriptome, a primer corresponding to the fragment is further designed and synthesized, PCR amplification is carried out, and after sequencing of an amplification product, the migratory locust Rab5 gene is obtained, and the sequence is shown as SEQ ID NO: 1. Furthermore, according to SEQ ID NO: 1, a corresponding primer is designed and synthesized, a gene segment with the nucleotide sequence shown in SEQ ID NO: 2 is obtained through PCR amplification, the gene segment is subjected to in-vitro transcription according to a kit specification to synthesize dsRNA, the dsRNA is injected into a migratory locust body cavity to specifically silence a target gene, migratory locust dies during ecdysis, and multiple experiments show that the fatality rate reaches 100%. Due to the specificity and the efficient fatality rate of the dsRNA, the dsRNA has important practical significance for migratory locust prevention and control, and a new way can be provided for migratory locust prevention and control.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a migratory locust Rab5 gene and its application in migratory locust control. Background technique [0002] Migratory locusts have a wide range of distribution due to their characteristics of migration, strong reproductive ability, and miscellaneous feeding habits. Once a locust plague breaks out, it will cause serious harm to food production. At present, the control of locust disasters is mainly chemical insecticides. The long-term use of chemical insecticides has brought about many problems such as the emergence of locust resistance, pesticide residues, food safety hazards, and environmental pollution. Therefore, it is imminent to develop new environmentally friendly and effective biopesticides. [0003] RNA interference (RNAi) refers to the highly conserved phenomenon of specific post-transcriptional gene silencing triggered by double-stranded RNA (double-stranded RNA, dsRNA) in ...

Claims

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Application Information

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IPC IPC(8): C12N15/55C12N15/10C12N15/113C12N15/11A01N63/60A01P7/04
Inventor 王艳丽范云鹤李涛史学凯张建珍
Owner SHANXI UNIV
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