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Chilo suppressalis SDR gene and encoded protein and application thereof, dsRNA and amplified primer pair and application thereof

A technology of gene coding and diploid borer, applied in the field of insect genetic engineering, can solve the problem of less research on SDR and achieve the effect of increasing mortality

Inactive Publication Date: 2019-08-20
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, the research on Short-chain dehydrogenases / reductases (SDR) in insects mainly focuses on its molecular mechanism and function, and there are few studies on the application of SDR.

Method used

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  • Chilo suppressalis SDR gene and encoded protein and application thereof, dsRNA and amplified primer pair and application thereof
  • Chilo suppressalis SDR gene and encoded protein and application thereof, dsRNA and amplified primer pair and application thereof
  • Chilo suppressalis SDR gene and encoded protein and application thereof, dsRNA and amplified primer pair and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Cloning and Analysis of Short-chain dehydrogenases / reductases(SDR) Gene of Chilo suppressalis

[0047] 1. Extraction of total RNA of C. borer: Weigh 20 mg of C. borer sample into a 1.5 ml enzyme-free tube, use disposable enzyme-free grinding rod and liquid nitrogen to grind thoroughly, and use Promega's SV total RNAisolation system to extract The kit extracts total RNA, and the detailed steps refer to the kit instruction.

[0048] 2. Synthesis of cDNA: The total RNA extracted in the above 1 was synthesized into a cDNA template using PrimeScriptTM RT MasterMix Reverse Transcription Kit of Treasure Bioengineering Dalian Co., Ltd. (refer to the kit manual for detailed steps).

[0049] 3. Primer design: use transcriptome sequencing to obtain the Short-chain dehydrogenases / reductases (SDR) gene nucleic acid sequence (see SEQ ID NO: 1), and design primers to verify the predicted open reading frame. Design and synthesize the following primers:

[0050] Upstream primer sequen...

Embodiment 2

[0059] dsRNA synthesis

[0060] 1. Preparation of dsRNA template:

[0061] According to the Short-chain dehydrogenases / reductases (SDR) gene sequence obtained in Example 1, the dsRNA region was predicted by siDirect version 2.0, and NCBI Primer-BLAST was used

[0062] ( https: / / www.ncbi.nlm.nih.gov / tools / primer-blast / index.cgi?id=0 LINK_LOC= Blast Home ) Design specific amplification primers (5'-ends plus appropriate restriction sites) for the amplification of the dsRNA fragment (SEQ ID No.3) of the HeatShock Proteins (HSPs) gene, the designed specific primers are as follows :

[0063] Upstream primer sequence ds SDR-F (SEQ ID No.4):

[0064] TGGAATTCCCGATAAT GCTAA CCAAGTCGATG

[0065] Downstream primer sequence ds SDR-R (SEQ ID No.5):

[0066] TGGAATTCGCCGTCGTAC TGCAGGA a.

[0067] Note: The underlined part series are EcoRI restriction sites

[0068] The above primers ds SDR-F and ds SDR-R were used for PCR amplification, and the PCR reaction system was referre...

Embodiment 3

[0082] The efficiency of gene silencing after feeding dsRNA of Short-chain dehydrogenases / reductases (SDR) gene and its effect on the growth and development of Chilo suppressalis.

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Abstract

The present invention provides a chilo suppressalis SDR gene and an encoded protein and an application thereof, dsRNA and an amplified primer pair and an application thereof, and belongs to the technical field of insect genetic engineering. The chilo suppressalis SDR gene has a nucleotide sequence shown in SEQ ID No.1. The provided chilo suppressalis SDR gene participates in growth and developmentprocesses of chilo suppressalis, deletion of the gene significantly increases mortality of larvae of the chilo suppressalis. The dsRNA of the chilo suppressalis SDR gene inhibits expression of the chilo suppressalis SDR gene.

Description

technical field [0001] The invention belongs to the technical field of insect genetic engineering, and in particular relates to an SDR gene of Chilo suppressalis and its coded protein and its application, dsRNA and its amplified primer pair and its application. Background technique [0002] Chilo suppressalis (Lepidoptera: Boreridae) is an important polyphagous agricultural pest, which mainly harms rice, wild rice stem and corn (Zhou Ruiqi et al., rice population and wild rice stem population of Chilo suppressalis are resistant to pesticides and Sensitivity comparison of Cry proteins [J]. Chinese Science and Technology Papers, 2014, 9(09): 1075-1079). In recent years, with the change of rice planting structure, the reform of farming system, the promotion of cultivation measures such as dense planting and high fertilizer, climate warming and other factors, the number of occurrences and the degree of damage of C. Rice production poses a serious threat (Srensen JG et al. Massr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53C12N9/04C12N15/113A01N57/16A01P7/04C12N15/82C12N15/11A01H5/00A01H6/46
CPCA01N57/16C12N9/0006C12N15/1137C12N15/8218C12N15/8286C12N2310/14C12Y101/01
Inventor 马伟华孙亚杰赵景陈浩林拥军
Owner HUAZHONG AGRI UNIV
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