CN112695045B - Lepidoptera insect Hpx12 gene and application thereof - Google Patents

Abstract

The invention provides a lepidoptera insectHpx12The invention provides a gene and application thereof, and aims to provide a lepidoptera insect innate immune response regulation geneHpx12(ii) a The second purpose is to provide the lepidopteran insect innate immune response regulation geneHpx12The cloning method of (4); the third purpose is to provide the lepidopteran insect innate immune response regulation geneHpx12The use of (1). The invention relates to lepidopteran insectsHpx12After the gene is inhibited and expressed, the innate immune response of the polypide can be inhibited, the susceptibility of the polypide to germs is increased, the silkworms are more easily infected with BmNPV viruses, and further, the death rate of lepidoptera insects is greatly increased. If inhibiting lepidopteran insectsHpx12Can realize the purpose of pest control. Has positive significance for solving the problems of ecological deterioration, pesticide resistance and the like caused by chemical pesticides.

Description

Lepidoptera insect Hpx12 gene and application thereof

Technical Field

The invention belongs to the technical field of genetic engineering, and particularly relates to a lepidoptera insectHpx12Genes and applications thereof.

Background

Lepidoptera includes two types of insects, i.e., moth and butterfly, and belongs to the subclass pteroidea and holomorphia. About 20 thousands of species are known worldwide, and about 8000 more species are known in china. This order is the 2 nd largest order of the Insecta, second only to the Coleoptera. The distribution range is very wide, and the tropical varieties are most abundant. Most kinds of larvae are harmful to various cultivated plants, and those with larger body form usually eat leaves or bore branches completely. Smaller patients often suffer from leaf curl, leaf ornamentation, scabbing, silking and netting, or drilling into plant tissues to eat. There are many important pests such as peach fruit borer, apple leaf roller, cotton bollworm, cabbage butterfly, diamond back moth and many lepidoptera granary worms such as indian meal moth, etc. In addition, the famous silkworms and tussah silkworms also belong to the order of insects. Silkworm is a lepidoptera seriation insect using mulberry leaves as food, and the silkworm passes through the development stages of eggs, larvae, pupae and imagoes which are completely different in morphology and physiological function in life.

Insects are the most flourishing species group on the earth land at present and are inexhaustible resource treasury of human beings. In recent years, the immunity of insects has attracted great attention in the aspects of basic and application research, and the research on genes, signals and other problems related to the immunity of insects through experiments has very important practical significance for pest control, insect benefiting and disease prevention, development and utilization of antibacterial substances, research on human immune mechanisms and the like.

The Hpx12 protein is a multifunctional protein, has a peroxidase conserved domain, has an integrin binding site, and can be combined with transmembrane receptor integrin, thereby realizing the adhesion between cells and between the cells and extracellular matrix and leading blood cells to play the roles of phagocytosis and coating. The Hpx12 protein has been studied less in Insecta, and the function of the protein has been studied less in Bombyx mori.

Disclosure of Invention

The present invention is based on at least one of the above technical problems, and the present invention relates to lepidopteran insectsHpx12The gene can inhibit the innate immune response of the insect body after being inhibited and expressed, the susceptibility of the insect body to germs is increased, the silkworm is more easily infected with BmNPV virus, and further the death rate of lepidoptera insects is greatly increased. If inhibiting lepidopteran insectsHpx12The expression of the gene can realize the purpose of pest control. For solving the problems caused by chemical pesticidesThe problems of ecological deterioration, pesticide resistance and the like have positive significance.

In view of this, the present invention provides a lepidopteran insectHpx12The gene(s) is (are),Hpx12the gene is lepidoptera insect innate immune response regulating gene, and the nucleotide sequence is SEQ ID No.1.

According to the second aspect of the invention, it proposesHpx12The gene expression inhibitor and/or activity inhibitor is applied to lepidoptera insects as a pest control target.

According to the third aspect of the invention, it proposesHpx12The gene coded protein has the amino acid sequence of SEQ ID No.2.

According to a fourth aspect of the present invention, there is providedHpx12Gene-encoded proteins inhibit lepidopteran insects in vivoHpx12The ds RNA of gene expression is used for reducing the innate immune response of lepidoptera insects and increasing the susceptibility of the lepidoptera insects to pathogenic bacteria.

Further, the ds RNA sequence is SEQ ID No.3.

Further, the sequence of the primer for amplifying the ds RNA is SEQ ID No. 6-SEQ ID No.7;

a forward primer: 5' -GTAATACGACTCACTATAGGGAGTTCGCTAT

GAATCGTTTTC -3’；SEQ ID No. 6；

Reverse primer: 5' -GTAATACGACTCACTATAGGGAGATTAGTTGCAGCTT

TAAAATCATTAA -3’；SEQ ID No. 7。

Further, the lepidopteran insect is a silkworm.

According to the fourth aspect of the invention, cloning rights are proposedHpx12A method of genetic engineering comprising the steps of:

(1) Mosquito according to the anophelesHpx12Protein sequence of gene (Gen Bank accession number (XP _ 311449), searching NCBI database to obtain homologous gene in silkwormHpx12And (4) sequencing. The sequence is used for designing gene cloning primers: SEQ ID No.4-SEQ ID No.5.

Synthesizing lepidoptera insect body cDNA: extracting lepidopterous insect RNA, and performing reverse transcription to obtain first-strand cDNA;

extracting the RNA of the whole silkworm, and performing reverse transcription to obtain a first chain c DNA;

（2）Hpx12PCR amplification of the genes: taking the lepidoptera insect body cDNA as a template, designing a primer according to the sequence of the lepidoptera insect body, carrying out PCR amplification, recovering and purifying a PCR amplification product, and sequencing.

According toHpx12Designing and synthesizing a specific primer by a gene sequence, carrying out PCR amplification by taking a first chain c DNA obtained by reverse transcription as a template, selecting a TransStart high-fidelity amplification enzyme reaction system, and ensuring that the total volume of the system is 50 mu L, wherein the specific primer comprises the following components: 200ng c DNA,5 × TransStart FastPfu reaction buffer 10 μ L,10m Md NTP 1 μ L,2U TransStart FastPfu DNA Polymerase,1 μ L of 1 μ M each of the forward and reverse primers, and water to 50 μ L. The PCR reaction is carried out on a Mastercycler pro amplification instrument, and the reaction program is as follows: 4 minutes at 94 ℃; 30 cycles of 94 ℃,30 seconds, 58 ℃,30 seconds, 72 ℃,1 minute; extension at 72 ℃ for 10 min; the PCR product was recovered and purified.

The purified product is connected with a vector and is connected with pEASY-Blunt Cloning Kit vectors through a Kit reaction, and a connection system and the process are as follows: uniformly mixing 4 mu L of the purified product and 1 mu L of pEASY-Blunt Cloning Kit, and carrying out water bath for 30 min at 25 ℃; transforming the connected vector into escherichia coli DH5a by heat shock, adding a liquid culture medium, performing shake culture, coating the cultured product on an LB (lysogeny broth) plate containing 100mg/L kanamycin for overnight culture, selecting bacterial colonies, performing bacterial liquid culture, extracting plasmids and performing PCR (polymerase chain reaction) detection. Screening positive clones, and sequencing the positive clones.

The invention firstly provides a lepidoptera insect innate immune response regulation geneHpx12Regulating gene for inhibiting lepidopteran insect innate immune responseHpx12The expression of the gene can reduce the innate immune response of lepidoptera insects, further increase the susceptibility of insect bodies to germs, and lead to the theory that the death rate of the lepidoptera insects is greatly increased, and has positive significance for solving the problems of ecological deterioration, insecticide resistance and the like caused by chemical insecticides.

Further, the primer sequence in the step (2) is SEQ ID No.4-SEQ ID No.5;

a forward primer: 5' -CGGGATCCGTTCGCTATGAATCGTTTTC-3’； SEQ ID No.4；

Reverse primer: 5' -GGGTTCGAATTAGTTGCAGCTTTAAAATCATTAA-3’；SEQ ID No.5。

Through the technical scheme, the invention provides a lepidoptera insectHpx12The invention relates to a gene, protein and application, wherein the invention uses homologous clone technique to obtain a lepidoptera insect innate immune response regulation gene from silkwormHpx12First proposed and utilizedHpx12Gene expressed protein, gene for suppressing lepidopteran insect innate immune response regulationHpx12Expression of (a): the innate immune response of the lepidoptera insects is reduced, so that the susceptibility of the insects to germs is increased, and the death rate of the lepidoptera insects is greatly increased.

Drawings

FIG. 1 shows the results of example 1 of the present inventionHpx12Agarose gel electrophoresis results of the genes;

wherein, M is DNA Marker, and 1 is target segment.

FIG. 2 shows the results of example 2 of the present inventionHpx12After the gene ds RNA is injected into the worm,Hpx12the expression level of the gene at different times.

FIG. 3 shows a schematic diagram of a display device according to embodiment 3 of the present inventionHpx12Number of nodes of silkworm larvae treated with BmNPV virus after injection of gene ds RNA into the body.

FIG. 4 shows a schematic diagram of a display device in example 4 of the present inventionHpx12Expression level of the antimicrobial peptide related gene 48h after injection of the gene ds RNA into the worm.

FIG. 5 shows a flowchart of example 5 of the present inventionHpx12The death rate of the silkworm larvae is 48h after the gene ds RNA is injected into the silkworm bodies.

Detailed Description

In order that the above objects, features and advantages of the present invention can be more clearly understood, the present invention will be described in further detail with reference to specific embodiments. It should be noted that the embodiments and features of the embodiments of the present invention may be combined with each other without conflict.

In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention, however, the present invention may be practiced in other ways than those specifically described herein, and therefore the scope of the present invention is not limited by the specific embodiments disclosed below.

Example 1

Hpx12Cloning of genes

(1) Mosquito according to the anophelesHpx12Protein sequence of gene (Gen Bank accession number (XP _ 311449), searching NCBI database to obtain homologous gene in silkwormHpx12And (4) sequencing.

Lepidoptera insect innate immune response regulating geneHpx12The nucleotide sequence of (A) is shown in SEQ ID NO.1.

Designing gene cloning primer ID No.4-SEQ ID No.5 by using the sequence;

a forward primer: 5' -CGGGATCCGTTCGCTATGAATCGTTTTC-3’； SEQ ID No.4；

Reverse primer: 5' -GGGTTCGAATTAGTTGCAGCTTTAAAATCATTAA-3’；SEQ ID No.5。

B. Extracting the RNA of the whole silkworm, and performing reverse transcription to obtain a first chain c DNA;

C. according toHpx12The gene sequence is obtained by performing PCR amplification by using primers SEQ ID No.4-SEQ ID No.5 and a first chain c DNA obtained by reverse transcription as a template, and a TransStart high fidelity amplification enzyme reaction system is selected, wherein the total volume of the system is 50 mu L, and the method comprises the following steps: 200ng c DNA,5 × TransStart FastPfu reaction buffer 10 μ L,10m Md NTP 1 μ L,2U TransStart FastPfu DNA Polymerase,1 μ L of 1 μ M each of the forward and reverse primers, and water to 50 μ L. The PCR reaction is carried out on a Mastercycler pro amplification instrument, and the reaction program is as follows: 4 minutes at 94 ℃; 30 cycles of 94 ℃,30 seconds, 58 ℃,30 seconds, 72 ℃,1 minute; extension at 72 ℃ for 10 min; the PCR product was recovered and purified, and the result is shown in FIG. 1, and the desired product gene was obtained as shown in FIG. 1.

The purified product is connected with a vector and is connected with pEASY-Blunt Cloning Kit vectors through a Kit reaction, and a connection system and the process are as follows: uniformly mixing 4 mu L of the purified product and 1 mu L of pEASY-Blunt Cloning Kit, and carrying out water bath for 30 min at 25 ℃; transforming the connected vector into escherichia coli DH5a by heat shock, adding a liquid culture medium, performing shake culture, coating the cultured product on an LB (lysogeny broth) plate containing 100mg/L kanamycin for overnight culture, selecting bacterial colonies, performing bacterial liquid culture, extracting plasmids and performing PCR (polymerase chain reaction) detection. Screening positive clones, and sequencing the positive clones.

Example 2

Lepidoptera insectsHpx12Synthesis and microinjection of Gene ds RNA

Lepidoptera insects obtained by cloning according to example 1Hpx12Full-length sequence SEQ ID NO.1 in gene, designHpx12Ds RNA of the gene is synthesized into ds RNA with a sequence of SEQ ID No.3 by an in vitro transcription reagent, and specifically, primers SEQ ID No. 6-SEQ ID No.7 are designed by a RiboMAX Large Scale RNA Production System-T7 kit;

a forward primer: 5' -GTAATACGACTCACTATAGGGAGTTCGCTAT

GAATCGTTTTC -3’；SEQ ID No. 6；

Reverse primer: 5' -GTAATACGACTACACTATAGGGAGATTAGTTGCAGCTT

TAAAATCATTAA -3’；SEQ ID No. 7。

Selecting silkworm larvae of 5 th 2 days old, uniform in size and consistent in health condition, setting an injected dsGFP control group and an injected dsHpx12 experimental group, anaesthetizing the silkworm larvae on ice before injection, then injecting ds RNA synthesized in vitro into a body cavity of the silkworm larvae by using a micro-injector, setting 3 biological repetitions by injecting 60 heads in each group, and feeding the silkworm larvae in a climatic chamber after injection.

Fluorescence quantitative PCR is used for detecting 6h, 12h, 24h and 48h after microinjectionHpx12As shown in FIG. 2, the expression and effect of the gene are the best interference effect in 24h, and the silkworm can be efficiently silenced in the case of the optimal interference effect in 24hHpx12The expression of the gene has better silencing effect of the target gene.

Example 3

Lepidoptera insectsHpx12The ds RNA of the gene can inhibit the innate immune response of the polypide and increase the susceptibility of the polypide to pathogenic bacteria

The surface disinfection is carried out on the 5 th-day and 3-day larva of the silkworm by using 75% ethanol. 2ml of BmNPV was injected into the blood chamber of the silkworms which had been injected with dsRNA by a microinjector. At 25 deg.CAfter standing for 2h, melanin and dark nodules were counted under a microscope at 50-fold magnification. Control insects were injected with only 2ml of BmNPV. Each test method was repeated three times as shown in fig. 3. As can be seen from FIG. 3, the number of nodules was significantly reduced in the ds RNA-injected treated group compared to the control group, indicating that lepidopteran insectsHpx12After the expression of the gene is inhibited, the immunity of the silkworm is greatly reduced.

The expression of the antibacterial peptide genes (Defension 1 gene and CeCA gene) at 24h after the microinjection of ds RNA is detected by utilizing fluorescent quantitative PCR, the result of the treatment group injected with ds RNA is shown in figure 4 compared with the control group, and as can be seen from figure 4, the expression level of the antibacterial peptide genes is obviously reduced, which indicates that lepidoptera insects are shownHpx12After the expression of the gene is inhibited, the immune response of the silkworm is greatly reduced.

Silkworm (Bombyx mori)Hpx12Feeding individuals with down-regulated gene expression to a feed containing 1.5X 10 ⁴ The results of bioassay on fresh mulberry leaves of BmNPV are shown in FIG. 5, and it can be seen from FIG. 5 thatHpx12The sensitivity of the silkworm with the down-regulated expression to BmNPV is obviously enhanced, and the mortality rate of the silkworm at 48h is obviously higher than that of the control.

Therefore, the design method for reducing the drug resistance of the pests on the basis of the molecular level has important significance for improving the pesticide effect of the biological pesticide and enhancing the pest control effect.

The above is only a preferred embodiment of the present invention, and is not intended to limit the present invention, and various modifications and changes will occur to those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention. .

Sequence listing

<110> Zhengzhou university of light industry

<120> lepidoptera insect Hpx12 gene and application thereof

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agattgttac ccgaagccga ctgtggagac gatgaaacct ctccatctgg agctccttta 300

aaatctgccc gggaagtcag gcagcgtatt ttacagacag ggaaggcatc agacctctca 360

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cgctgcttga atctcactcg accgaaatcg ttccagactt atggctgcct tgctgactcg 600

aatgtagaca gaatcgaatt cacaacaccg ttatttgatt tatcgactat ctaccgatcg 660

accgaaccgg ctccagaata cagaacatac agtggcggat tactgatgac ccaagaggcc 720

gatggcacca tcttcccccc acaagaagga ccgcacagca ataaatgctt gcaaaacgac 780

gcgtctaatg gagaaacgaa atgttttggc cctgtttcaa catcgattct gccggtcaca 840

ttgttggtcg tgtggtggtg gagactacac aataagatcg ccaaagaact aaacgaaatc 900

aatcctcatt gggatgacga aacattgttc caaactgcaa gggacattaa catcgcaata 960

acaaaccaat ttgtttacta cgaattgctg ccaactctat tcggtgaaga attctgtttg 1020

aaaaatgagc tgattcactc tgaatctgga cacagagacc tttacgatga gtctatacct 1080

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ttgaaacttt tcgacgagga tttcaagtac gtgggcaaaa aaatggtaac agacctctct 1200

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Met Leu Thr Ser Leu Val Pro Leu Ala Pro Leu Leu Leu Thr Ala Gly

1 5 10 15

Val Ala Thr Gly Ser Pro Ser Gly Leu Pro Ala Thr Ala Gly Leu Ile

20 25 30

Gly Thr Pro Ile Leu Gly Gly Ala Leu Ala Ala Cys Thr Ala Ala Ile

35 40 45

Gly Leu Pro Cys Ala Pro Ala Gly Ala Ala Ala Leu Ala Gly Ser Cys

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Ser Ala Pro Ala Thr Pro Ser Ala Gly Thr Pro His Thr Pro Val Ile

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Ala Leu Leu Pro Gly Ala Ala Cys Gly Ala Ala Gly Thr Ser Pro Ser

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Gly Ala Pro Leu Leu Ser Ala Ala Gly Val Ala Gly Ala Ile Leu Gly

100 105 110

Thr Gly Leu Ala Ser Ala Leu Ser Thr Thr Gly Leu Leu Ala Ile Ile

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Ser Thr Ile Ile Pro Ala Ala Leu Gly Ser Ile His Ala Ser Val Ala

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Leu Leu Thr Gly Thr Thr Ala Cys Cys Thr Ala Gly Gly Leu Ser Ala

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Thr Met Cys Thr Pro Ile Ala Ile Pro Gly Ala Ala Pro Val His Ala

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Pro Ser Gly Ile Ala Cys Leu Ala Leu Thr Ala Pro Leu Ser Pro Gly

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Thr Thr Gly Cys Leu Ala Ala Ser Ala Val Ala Ala Ile Gly Pro Thr

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Thr Pro Leu Pro Ala Leu Ser Thr Ile Thr Ala Ser Thr Gly Pro Ala

210 215 220

Pro Gly Thr Ala Thr Thr Ser Gly Gly Leu Leu Met Thr Gly Gly Ala

225 230 235 240

Ala Gly Thr Ile Pro Pro Pro Gly Gly Gly Pro His Ser Ala Leu Cys

245 250 255

Leu Gly Ala Ala Ala Ser Ala Gly Gly Thr Leu Cys Pro Gly Pro Val

260 265 270

Ser Thr Ser Ile Leu Pro Val Thr Leu Leu Val Val Thr Thr Thr Ala

275 280 285

Leu His Ala Leu Ile Ala Leu Gly Leu Ala Gly Ile Ala Pro His Thr

290 295 300

Ala Ala Gly Thr Leu Pro Gly Thr Ala Ala Ala Ile Ala Ile Ala Ile

305 310 315 320

Thr Ala Gly Pro Val Thr Thr Gly Leu Leu Pro Thr Leu Pro Gly Gly

325 330 335

Gly Pro Cys Leu Leu Ala Gly Leu Ile His Ser Gly Ser Gly His Ala

340 345 350

Ala Leu Thr Ala Gly Ser Ile Pro Ala Thr Thr Leu Gly Thr Thr Leu

355 360 365

Ala Leu Ala Thr Pro His Leu Val Ser Gly Gly Ala Leu Leu Leu Pro

370 375 380

Ala Gly Ala Pro Leu Thr Val Gly Leu Leu Met Val Thr Ala Leu Ser

385 390 395 400

Leu His Thr Ala Pro Leu Met Ala Ala Leu Ala Leu Ala Leu Met Thr

405 410 415

Ala Gly Thr Thr Thr Gly Ala Gly Gly Ala Ala Ala Ala Ala Val Ala

420 425 430

Pro Ala Ile Cys Ala Leu Gly Leu Gly Ile Met Gly Leu Ala Ser Ala

435 440 445

Leu Thr Ala Ala Ala Leu Ala Leu Ala Gly Leu Pro Leu Ile Pro Pro

450 455 460

Thr Val Ala Thr Val Leu Leu Cys His Ala Val Gly Ile Leu Thr Thr

465 470 475 480

Leu Ala Met Leu Leu Pro Ile Ala Met Ala Ala Ile Gly Ser Leu Gly

485 490 495

Gly Ile Thr Gly Thr Pro Gly Ala Val Gly Leu Leu Ala Gly Ile Thr

500 505 510

Ile Gly Ala Pro Met Gly Gly Gly Thr Val Pro Pro Thr Ala Ala Cys

515 520 525

Ile Ile Ala Leu Gly Leu Ser Leu Thr Met Leu Ala Ala Ala His Thr

530 535 540

Thr Gly Ala Ser Ala Ala Pro Thr Ala Pro Ala Val Ala Gly Leu Ala

545 550 555 560

Gly Ile Ala Leu Ala Thr Val Ala Gly Pro Leu Cys Gly Val Gly Ala

565 570 575

Gly Val Gly Ala Ala Gly Pro Leu Cys Gly Val Gly Ala Gly Val Gly

580 585 590

Ala Ile Gly Ala Gly Ala Leu Leu Ala Val Thr Ala Ser Ala Pro Leu

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Val Ser Cys Gly Gly Ile Pro Ala Thr Ala Leu Ala Ala Thr Leu Gly

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Ser Leu

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ggagacgatg aaacctctcc atctggagct cctttaaaat ctgcccggga agtcaggcag 300

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tttggccctg tttcaacatc gattctgccg gtcacattgt tggtcgtgtg gtggtggaga 840

ctacacaata agatcgccaa agaactaaac gaaatcaatc ctcattggga tgacgaaaca 900

ttgttccaaa ctgcaaggga cattaacatc gcaataacaa accaatttgt ttactacgaa 960

ttgctgccaa ctctattcgg tgaagaattc tgtttgaaaa atgagctgat tcactctgaa 1020

tctggacaca gagaccttta cgatgagtct atacctgcta cttatctcga atactacctc 1080

gctttgagat ggttccactt ggtttcagag ggtgatttga aacttttcga cgaggatttc 1140

aagtacgtgg gcaaaaaaat ggtaacagac ctctctttac atactgattt cttgatgaga 1200

gataagaact tggcaaaaat gacacgagga acttactatc aggctggagg agataatgat 1260

agagctgtag acccggccat ttgtgataag ggattaggaa taatgcaaaa agcatccgac 1320

cttacagcag cggatttgag gaaaaatcag cttttcaaaa ttccacctta cgtcgactac 1380

gtaaaactgt gccatgatgt cgaaattaaa acatggaaag acatgttgaa attcattgac 1440

atggatcgaa ttgaaagtct gcaagaaatt tacgaaaccc ctggtgacgt ggaactcctg 1500

gcaggaatct ggattgaaag acctatggag ggaggttacg ttcctccaac cgcagcttgt 1560

atcataaaca aacaactgtc tctcactatg aaagctgaca gacattggta tgagagatcg 1620

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ggcttccttt gcgaagttgg tgacggtgta gagagaatcc aacgggaggc acttaaaaga 1740

gtgactgctt cgaatccatt ggtcagctgc caagaaatcc cgagatggaa tctcgctgct 1800

tggaaagaat ctaagtgaga ttaatgattt taaagctgca actaatctcc ctatagtgag 1860

tcgtattac 1869

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<212> DNA

<213> Artificial Sequence (Artificial Sequence)

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cgggatccgt tcgctatgaa tcgttttc 28

<210> 5

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<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 5

gggttcgaat tagttgcagc tttaaaatca ttaa 34

<210> 6

<211> 44

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 6

gtaatacgac tcactatagg gagagttcgc tatgaatcgt tttc 44

<210> 7

<211> 49

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 7

gtaatacgac tcactatagg gagattagtt gcagctttaa aatcattaa 49

Claims (3)

1. A kind ofHpx12The gene expression inhibitor and/or activity inhibitor is applied to pest control; characterized in that the aboveHpx12The gene is the silkworm innate immune response regulation gene, and the nucleotide sequence is SEQ ID No.1; the pests are silkworms.

2. Inhibiting the said in vivo of silkwormHpx12The application of the gene expression ds RNA in reducing the innate immune response of silkworms and increasing the susceptibility of the silkworms to germs is characterized in that the sequence of the ds RNA is SEQ ID No.3.

3. The use according to claim 2, wherein the primer sequence for amplifying the ds RNA is SEQ ID No.6 to SEQ ID No.7;

a forward primer: 5' -GTAATACGACTCACTATAGGGAGTTCGCTAT

GAATCGTTTTC -3’；SEQ ID No. 6；

Reverse primer: 5' -GTAATACGACTCACTATAGGGAGATTAGTTGCAGCTT

TAAAATCATTAA -3’；SEQ ID No. 7。

Images