CN111394371A - Migratory locust V-ATPase-V1 structural domain gene and application of dsRNA thereof in pest control - Google Patents

Migratory locust V-ATPase-V1 structural domain gene and application of dsRNA thereof in pest control Download PDF

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CN111394371A
CN111394371A CN202010249056.6A CN202010249056A CN111394371A CN 111394371 A CN111394371 A CN 111394371A CN 202010249056 A CN202010249056 A CN 202010249056A CN 111394371 A CN111394371 A CN 111394371A
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刘晓健
梁晓宇
史学凯
张建珍
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Abstract

The invention discloses a migratory locust V-ATPase-V1 structural domain gene and application of dsRNA thereof in pest control, wherein the migratory locust V-ATPase-V1 structural domain gene comprises migratory locust V-ATPase-A, V-ATPase-B, V-ATPase-C, V-ATPase-D, V-ATPase-E, V-ATPase-F and V-ATPase-G genes, and the V-ATPase-A to V-ATPase-G genes are obtained from a migratory locust transcription group database by a bioinformatics method, and the full-length cDNA sequences of the genes are obtained after cloning and sequencing the genes respectively; based on the gene sequences, V-ATPase-A to V-ATPase-G gene fragments are respectively obtained by a PCR method, corresponding dsRNA is synthesized, and the synthesized dsRNA can be respectively injected into a locusta migratoria coelomis and then can specifically silence a target gene, so that the locusta migratoria is died due to the fact that growth and development of the locusta migratoria are blocked, and the lethality rate reaches 60.3% -93.7%. Due to the specificity and the high-efficiency lethality rate, the invention has important significance for pest control and provides a new way for pest control.

Description

Migratory locust V-ATPase-V1 structural domain gene and application of dsRNA thereof in pest control
Technical Field
The invention relates to the field of biotechnology and agricultural pest control, in particular to a migratory locust V-ATPase-V1 structural domain gene and application of dsRNA thereof in pest control.
Background
The migratory locust L austria mightaria is an intercontinental agricultural pest which is mainly distributed in four continents of Asia, Europe, Africa and Australia and has the characteristics of outbreak, cluster and migration, once the migratory locust occurs, the migratory locust has wide related range and fierce tendency, and serious disasters are caused.
RNA interference (RNAi), a phenomenon of specific post-transcriptional gene silencing caused by double-stranded RNA molecules, has been ascending at the technology front since the acquisition of the nobel prize in 2006. RNAi is not only a powerful tool for studying gene function, but also has great potential in pest control. Pest control by RNAi has the following characteristics: 1) the insecticidal has specificity and no killing effect on non-target organisms; 2) RNA is easy to degrade in nature and has no residue; 3) is nontoxic and harmless to the environment and is relatively safe. Currently, the RNAi technology has been internationally referred to as fourth generation pesticide core technology. RNAi transgenic maize (MON87411) expressing dsRNA of the maize root leaf beetle Snf7 gene, developed by Monsanto, USA, has been approved by the United States Department of Agriculture (USDA) and the Environmental Protection Agency (EPA), and is about to be commercially used. Experiments show that the transgenic corn can effectively kill corn rootworm and has no obvious toxic effect on non-target organisms such as bees and the like. Screening and identifying genes which are crucial to the growth and development of pests are key links for developing novel pesticides by applying RNAi technology, because not all dsRNA can effectively silence the expression of target genes, nor all dsRNA can kill pests.
V-ATPase(Vacuolar-type ATPase), an H+The channel protein is widely present on membranes of various organelles such as endosomes, lysosomes and the like of prokaryotes and eukaryotes and plasma membranes of some special cells, regulates the pH of the internal and external environments of the cells and is important for maintaining normal life activities of organisms. The V-ATPase is a highly conserved complex enzyme, and is formed by assembling an extracellular V1 domain and a transmembrane V0 domain, wherein the V1 domain consists of a A, B, C, D, E, F, G, H large subunit and is responsible for the hydrolysis of ATP; the V0 domain consists of a, c, c ', d, e (c' is present in yeast cells), and functions as a proton transporter. At present, no relevant research aiming at the V-ATPase-V1 structural domain gene (V-ATPase-A, V-ATPase-B, V-ATPase-C, V-ATPase-D, V-ATPase-E, V-ATPase-F and V-ATPase-G) of locusta migratoria is found. The invention proves that 60.3% -93.7% of lethality occurs after dsRNA of the V-ATPase-V1 structural domain gene of the migratory locust is injected to 3-year-old nymphs respectively for the first time. Therefore, the V-ATPase-V1 structural domain gene can be used as a high lethal target gene for RNAi-based pest control technology.
Disclosure of Invention
Aiming at the current state of the prior art, the invention aims to provide a migratory locust V-ATPase-V1 structural domain gene, a gene fragment and application of dsRNA thereof in lethal migratory locust.
The invention provides a locusta migratoria V-ATPase-V1 structural domain gene, which comprises the following genes:
the nucleotide sequence of the migratory locust V-ATPase-A gene is SEQ ID NO: 1, the length of the nucleotide is 1848 bp; the amino acid sequence is SEQ ID NO: 2, the sequence length is 615 amino acids;
the nucleotide sequence of the locust V-ATPase-B gene is SEQ ID NO: 6, the length of the nucleotide is 1503 bp; the amino acid sequence is SEQ ID NO: 7, the sequence length is 500 amino acids;
the nucleotide sequence of the migratory locust V-ATPase-C gene is SEQ ID NO: 11, the length of the nucleotide is 1158 bp; the amino acid sequence is SEQ ID NO: 12, the sequence length is 385 amino acids;
the nucleotide sequence of the migratory locust V-ATPase-D gene is SEQ ID NO: 16, the length of the nucleotide is 753 bp; the amino acid sequence is SEQ ID NO: 17, the sequence length is 250 amino acids;
the nucleotide sequence of the migratory locust V-ATPase-E gene is SEQ ID NO: 21, the length of the nucleotide is 684 bp; the amino acid sequence is SEQ ID NO: 22, the sequence length is 227 amino acids;
the nucleotide sequence of the migratory locust V-ATPase-F gene is SEQ ID NO: 26, the length of the nucleotide is 372 bp; the amino acid sequence is SEQ ID NO: 27, the sequence length is 123 amino acids;
the nucleotide sequence of the migratory locust V-ATPase-F gene is SEQ ID NO: 31, the length of the nucleotide is 357 bp; the amino acid sequence is SEQ ID NO: 32, the sequence length is 118 amino acids.
The invention provides a locusta migratoria V-ATPase-V1 structural domain gene fragment, which comprises the following gene fragments:
the nucleotide sequence of the V-ATPase-A gene fragment is SEQ ID NO: 3;
the nucleotide sequence of the V-ATPase-B gene fragment is SEQ ID NO: 8;
the nucleotide sequence of the V-ATPase-C gene fragment is SEQ ID NO: 13;
the nucleotide sequence of the V-ATPase-D gene fragment is SEQ ID NO: 18;
the nucleotide sequence of the V-ATPase-E gene fragment is SEQ ID NO: 23;
the nucleotide sequence of the V-ATPase-F gene fragment is SEQ ID NO: 28;
the nucleotide sequence of the V-ATPase-G gene fragment is SEQ ID NO: 33.
the dsRNA of the V-ATPase-V1 structural domain gene of migratory locust provided by the invention comprises the following dsRNA:
the locust migratory V-ATPase-A gene fragment SEQ ID NO: 3 a synthetic dsRNA;
the locust migratory V-ATPase-B gene fragment SEQ ID NO: 8 synthetic dsRNA;
the locust-migratory V-ATPase-C gene fragment SEQ ID NO: 13 synthetic dsRNA;
the locust migratory V-ATPase-D gene fragment SEQ ID NO: 18 synthetic dsRNA;
the locust migratory V-ATPase-E gene fragment SEQ ID NO: 23 synthetic dsRNA;
the locust migratory V-ATPase-F gene fragment SEQ ID NO: 28 synthetic dsRNA;
the locust migratory V-ATPase-G gene fragment SEQ ID NO: 33 synthetic dsRNA.
The invention provides an application of migratory locust V-ATPase-V1 structural domain gene dsRNA in pest control, based on migratory locust V-ATPase-A, V-ATPase-B, V-ATPase-C, V-ATPase-D, V-ATPase-E, V-ATPase-F and V-ATPase-G genes contained in a migratory locust V-ATPase-V1 structural domain, for each gene, an upstream primer and a downstream primer containing a T7 promoter aiming at the gene are designed through primer premier 5.0 software, and a DNA template with two ends being T7 promoters is obtained through PCR amplification. According to HiScribeTMT7 High Yield RNA Synthesis Kit(NEW ENGLAND
Figure BDA0002433412330000031
Inc.) kit demonstrates in vitro transcription synthesis of the corresponding dsRNA. Each synthetic dsRNA was injected into the body cavity of a 3-year migratory locust via a microinjector. The results show that:
after the dsRNA is injected, the mRNA expression of the migratory locust V-ATPase-A gene is obviously reduced, and the migratory locust has growth and development resistance and leads to 79.7 percent of death.
After the dsRNA is injected, the mRNA expression of the migratory locust V-ATPase-B gene is obviously reduced, and the migratory locust has the defects of growth and development resistance and 85.7 percent of death.
After the dsRNA is injected, the mRNA expression of the V-ATPase-C gene of the migratory locust is obviously reduced, and the migratory locust has growth and development resistance and causes 69.8 percent of death.
After the dsRNA is injected, the mRNA expression of the migratory locust V-ATPase-D gene is obviously reduced, and the migratory locust has growth and development resistance and causes 90.4 percent of death.
After the dsRNA is injected, the mRNA expression of the migratory locust V-ATPase-E gene is obviously reduced, and the migratory locust has the defects of hindered growth and development and causes 93.7 percent of death.
After the dsRNA is injected, the mRNA expression of the migratory locust V-ATPase-F gene is obviously reduced, and the migratory locust has growth and development resistance and leads to 77.8 percent of death.
After the dsRNA is injected, the mRNA expression of the V-ATPase-G gene of the migratory locust is obviously reduced, and the migratory locust has growth and development resistance and causes 60.3 percent of death.
Wherein, in the synthesis of dsRNA of locusta migratoria V-ATPase-V1 structural domain gene, based on different genes, an upstream primer and a downstream primer are designed as follows:
the upstream primer and the downstream primer of the migratory locust V-ATPase-A gene are respectively as follows: SEQ ID NO: 4 and SEQ ID NO: 5;
the upstream primer and the downstream primer of the migratory locust V-ATPase-B gene are respectively as follows: SEQ ID NO: 9 and SEQ ID NO: 10;
the upstream primer and the downstream primer of the migratory locust V-ATPase-C gene are respectively as follows: SEQ ID NO: 14 and SEQ ID NO: 15;
the upstream primer and the downstream primer of the migratory locust V-ATPase-D gene are respectively as follows: SEQ ID NO: 19 and SEQ ID NO: 20;
the upstream primer and the downstream primer of the migratory locust V-ATPase-E gene are respectively as follows: SEQ ID NO: 24 and SEQ ID NO: 25;
the upstream primer and the downstream primer of the migratory locust V-ATPase-F gene are respectively as follows: SEQ ID NO: 29 and SEQ ID NO: 30, of a nitrogen-containing gas;
the upstream primer and the downstream primer of the migratory locust V-ATPase-G gene are respectively as follows: SEQ ID NO: 34 and SEQ ID NO: 35.
compared with the prior art, the invention has the following beneficial effects:
1. because of the specificity and high-efficiency lethality of dsRNA synthesized by the gene fragment of the V-ATPase-V1 domain of the migratory locust, the dsRNA has important practical significance for pest control and can provide a new way for pest control.
2. The dsRNA of the invention is easy to synthesize, is convenient to prepare into a spray type pesticide or bait, and has better popularization and application values.
Drawings
FIG. 1 shows the effect of dsRNA (ds L mV-ATPase-A) injection of V-ATPase-A gene on mRNA expression of V-ATPase-A gene after 3 rd migratory locust injection, dsGFP as control, ds L mV-ATPase-A as treatment group, and β -actin as internal reference gene, where P is < 0.001.
FIG. 2 shows the effect of dsRNA (ds L mV-ATPase-A) injected with V-ATPase-A gene on growth and development of 3-year migratory locusts, the left side is a control injected with dsGFP, the right side is a treatment group injected with dsRNA synthesized by SEQ ID NO. 3, namely ds L mV-ATPase-A, wherein 1 is the ridge line of the 3-year migratory locusts of the treatment group, the 4-year migratory locusts of the treatment group, the dorsal fissure is opened and the dorsal arch is formed and the locusts die, 2 is the death of the treatment group 1-2 days after the 3-year migratory locusts of the treatment group, 3 is the death of the treatment group 1-2 days before the 5-year migratory of the 4-year migratory locusts of the treatment group, and 4 is the death of the treatment group locust.
FIG. 3 shows the effect of dsRNA (ds L mV-ATPase-B) injection of the V-ATPase-B gene on mRNA expression of the V-ATPase-B gene after 3 rd migratory locust injection with dsGFP as a control, ds L mV-ATPase-B as a treatment group, and β -actin as an internal reference gene, where P < 0.01.
FIG. 4 shows the effect of dsRNA (ds L mV-ATPase-B) injected with V-ATPase-B gene on growth and development of 3-year migratory locusts, the left side is a control injected with dsGFP, the right side is a treatment group injected with dsRNA synthesized by SEQ ID NO. 8, namely ds L mV-ATPase-B, wherein 1 is the death 1-2 days before the 3-year sloughing of the treatment group migratory locusts at 4-year sloughing, 2 is the death of the old epidermic compact inclusion at 3-year sloughing of the treatment group migratory at 4-year sloughing, and 3 is the death of the treatment group migratory locusts at 1-2 days after the 3-year sloughing.
FIG. 5 shows the effect of dsRNA (ds L mV-ATPase-C) injection of the V-ATPase-C gene on mRNA expression of the V-ATPase-C gene after a 3-year migratory locust was reached, dsGFP was used as a control, ds L mV-ATPase-C was used as a treatment group, and β -actin was used as an internal reference gene, where P < 0.001.
FIG. 6 shows the effect of dsRNA injection of V-ATPase-C gene (ds L mV-ATPase-C) on growth and development of 3-year migratory locust after injection, the left side shows a control injected with dsGFP, the right side shows a treatment group injected with dsRNA synthesized by SEQ ID NO: 13, i.e. ds L mV-ATPase-C, wherein 1 shows that the ridge line of the 3-year migratory locust of the treatment group breaks and the back bows and dies at the 4-year slough, and 2 shows that the locust of the treatment group dies 1-2 days after the 4-year slough of the 3-year migratory locust of the treatment group is 4-year slough.
FIG. 7 shows the effect of dsRNA (ds L mV-ATPase-D) injection of the V-ATPase-D gene on mRNA expression of the V-ATPase-D gene after 3 rd migratory locust injection with dsGFP as a control, ds L mV-ATPase-C as a treatment group, and β -actin as an internal reference gene, where P < 0.01.
FIG. 8 shows the effect of dsRNA (ds L mV-ATPase-D) injection of V-ATPase-D gene on growth and development of 3-year migratory locust after injection, the left side shows a control injected with dsGFP, the right side shows a treatment group injected with dsRNA synthesized by SEQ ID NO. 18, namely ds L mV-ATPase-D, wherein 1 shows death 1-2 days before the 3-year slough of the treatment group migratory locust is 4-year slough, 2 shows that the old epidermal death cannot be successfully sloughed off when the 3-year migratory locust is 4-year slough of the treatment group migratory locust, and 3 shows death 1-2 days after the 3-year slough is 4-year slough of the treatment group migratory locust.
FIG. 9 shows the effect of dsRNA (ds L mV-ATPase-E) injection of the V-ATPase-E gene on mRNA expression of the V-ATPase-E gene after 3 rd migratory locust injection with dsGFP as a control, ds L mV-ATPase-E as a treatment group, and β -actin as an internal reference gene, where P < 0.001.
FIG. 10 shows the effect of dsRNA injection of V-ATPase-E gene (ds L mV-ATPase-E) on growth and development of migratory locust at 3 years, the left side shows control injection of dsGFP, the right side shows treatment group injection of dsRNA synthesized by SEQ ID NO. 23, i.e. ds L mV-ATPase-E, wherein 1 shows death 1-2 days before 3-year ghost of migratory locust at 4-year, and 2 shows ridge line open dorsal arch and death at 4-year ghost of migratory locust at 3-year ghost at 2.
FIG. 11 shows the effect of dsRNA (ds L mV-ATPase-F) injection of the V-ATPase-F gene on mRNA expression of the V-ATPase-F gene after 3 rd migratory locust injection with dsGFP as a control, ds L mV-ATPase-F as a treatment group, and β -actin as an internal reference gene, where P < 0.01.
FIG. 12 shows the effect of dsRNA (ds L mV-ATPase-F) injection of V-ATPase-F gene on growth and development of 3-year migratory locust, the left side shows control injection of dsGFP, the right side shows treatment group injection of dsRNA synthesized by SEQ ID NO: 28, i.e. ds L mV-ATPase-F, wherein 1 shows that the 3-year migratory locust in the treatment group fails to successfully slough off the old epidermis and dies at 4-year, and 2 shows that the migratory locust in the treatment group dies 1-2 days after 3-year sloughing and 4-year.
FIG. 13 shows the effect of dsRNA (ds L mV-ATPase-G) injection of the V-ATPase-G gene on mRNA expression of the V-ATPase-G gene after 3 rd migratory locust injection with dsGFP as a control, ds L mV-ATPase-G as a treatment group, and β -actin as an internal reference gene, where P < 0.001.
FIG. 14 shows the effect of dsRNA (ds L mV-ATPase-G) injection of V-ATPase-G gene on growth and development of migratory locust at 3 years, the left side shows control injection of dsGFP, the right side shows treatment group injection of dsRNA synthesized by SEQ ID NO. 33, i.e. ds L mV-ATPase-G, wherein 1 shows death 1-2 days before 3-year ghost of migratory locust at 4-year, and 2 shows that old epidermal death cannot be successfully sloughed off when 3-year ghost of migratory locust at 4-year old.
Detailed Description
The technical scheme of the invention is further explained by the specific embodiment in combination with the attached drawings. It should be understood by those skilled in the art that the specific embodiments are only for the understanding of the present invention and should not be construed as the specific limitations of the present invention.
The locusta migratoria V-ATPase-V1 structural domain gene comprises the following genes:
the nucleotide sequence of the migratory locust V-ATPase-A gene is SEQ ID NO: 1;
the nucleotide sequence of the migratory locust V-ATPase-B gene is SEQ ID NO: 6;
the nucleotide sequence of the migratory locust V-ATPase-C gene is SEQ ID NO: 11;
the nucleotide sequence of the migratory locust V-ATPase-D gene is SEQ ID NO: 16;
the nucleotide sequence of the migratory locust V-ATPase-E gene is SEQ ID NO: 21;
the nucleotide sequence of the migratory locust V-ATPase-F gene is SEQ ID NO: 26;
the nucleotide sequence of the migratory locust V-ATPase-G gene is SEQ ID NO: 31.
example 1: obtaining of each gene and gene fragment in V-ATPase-V1 structural domain of migratory locust
1) Acquisition of each gene in V-ATPase-V1 domain of migratory locust
The locusta migratoria V-ATPase-V1 structural domain genes comprise 7 genes of locusta migratoria V-ATPase-A, locusta migratoria V-ATPase-B, locusta migratoria V-ATPase-C, locusta migratoria V-ATPase-D, locusta migratoria V-ATPase-E, locusta migratoria V-ATPase-F and locusta migratoria V-ATPase-G.
On the basis of a transcriptome database of the migratory locust, searching migratory locust V-ATPase-A, migratory locust V-ATPase-B, migratory locust V-ATPase-C, migratory locust V-ATPase-D, migratory locust V-ATPase-E, migratory locust V-ATPase-F and migratory locust V-ATPase-G genes by a bioinformatics method to obtain a migratory locust V-ATPase-A gene (L mV-ATPase-A) sequence, a migratory locust V-ATPase-B gene (L mV-ATPase-B) sequence, a migratory locust V-ATPase-C gene (L mV-ATPase-C) sequence, a migratory locust V-ATPase-D gene (L mV-ATPase-D) sequence, a locust V-ATPase-E gene (L-ATPase-E) sequence, a migratory locust V-ATPase-D gene (ATPase-G-29 mV-ATPase-G) sequence.
Using primer premier 5.0 software to respectively design respective upstream and downstream primers for verifying each sequence obtained above, synthesizing the primers by biological engineering (Shanghai) corporation, selecting 3-year-old locusta migratoria 4 heads which are healthy in growth and identical in size and sex, quickly dissecting the body wall under a body type microscope, freezing the body wall, extracting RNA according to TaKaRaRNAiso Plus kit, using Reverse Transcriptase M-M L V (RNase H-) reagent according to instructions to reversely transcribe the extracted RNA into first-strand cDNA, using the cDNA as a template, combining the designed upstream and downstream primers of each gene, amplifying L mV-ATPase-A gene cDNA full-length sequence by PCR, L-ATPase-B gene cDNA full-length sequence, L mV-ATPase-C gene cDNA full-length sequence, L mV-ATPase-D gene cDNA sequence, 2mV cDNA-ATPase-E gene cDNA sequencing, separately cloning the cDNA-5 ATPase cDNA full-length sequence into the cDNA sequence of each gene, and respectively cloning the cDNA sequence of Escherichia coli genetic engineering (separately), and obtaining the result of the cDNA sequence obtained by cloning:
the V-ATPase-A gene sequence of locusta migratoria is SEQ ID NO: 1, the length of the nucleotide is 1848 bp;
the V-ATPase-B gene sequence of locusta migratoria is SEQ ID NO: 6, the length of the nucleotide is 1503 bp;
the V-ATPase-C gene sequence of locusta migratoria is SEQ ID NO: 11, the length of the nucleotide is 1158 bp;
the V-ATPase-D gene sequence of locusta migratoria is SEQ ID NO: 16, the length of the nucleotide is 753 bp;
the V-ATPase-E gene sequence of locusta migratoria is SEQ ID NO: 21, the length of the nucleotide is 684 bp;
the V-ATPase-F gene sequence of locusta migratoria is SEQ ID NO: 26, the length of the nucleotide is 372 bp;
the V-ATPase-G gene sequence of locusta migratoria is SEQ ID NO: 31, the nucleotide length is 357 bp.
2) Amino acid sequence analysis of each gene in V-ATPase-V1 domain of migratory locust
The nucleotide sequences of the obtained L mV-ATPase-A, L mV-ATPase-B, L mV-ATPase-C, L mV-ATPase-D, L mV-ATPase-E, L mV-ATPase-F and L mV-ATPase-G genes are translated by ExPasy online software, and the open reading frame coding, molecular weight and isoelectric point of each gene are predicted, and the results are as follows:
the open reading frame of L mV-ATPase-A gene encodes 615 amino acids, the molecular weight is 58KDa, and the isoelectric point is 5.11;
the open reading frame of L mV-ATPase-B gene encodes 500 amino acids, the molecular weight is 55.6KDa, and the isoelectric point is 5.33;
the open reading frame of the L mV-ATPase-C gene codes 385 amino acids, the molecular weight is 44.4KDa, and the isoelectric point is 8.16;
the open reading frame of L mV-ATPase-D gene encodes 250 amino acids, the molecular weight is 28KDa, and the isoelectric point is 9.63;
the open reading frame of the L mV-ATPase-E gene codes 227 amino acids, the molecular weight is 28KDa, and the isoelectric point is 8.51;
the open reading frame of the L mV-ATPase-F gene codes 123 amino acids, the molecular weight is 13.8KDa, and the isoelectric point is 5.92;
the open reading frame of the L mV-ATPase-G gene codes 118 amino acids, the molecular weight is 14KDa, and the isoelectric point is 9.66.
3) Obtaining of each gene fragment in V-ATPase-V1 domain of migratory locust
According to SEQ ID NO: 1, and a primer premier 5.0 software is adopted to design a specific primer, wherein the sequence of an upstream primer is SEQ ID NO: 4, the sequence of the downstream primer is SEQ ID NO: 5, all primers were synthesized by Biotechnology engineering (Shanghai) Inc. The method comprises the following steps of: 1 as a template, carrying out PCR amplification to obtain a migratory locust V-ATPase-A gene fragment, purifying and cloning the obtained product, transforming the purified product into escherichia coli, and sending the escherichia coli to a company Limited in Biotechnology engineering (Shanghai) for sequencing, wherein the sequence of the escherichia coli is SEQ ID NO: 3.
according to SEQ ID NO: 6, designing a specific primer by using primer premier 5.0 software, wherein the sequence of an upstream primer is SEQ ID NO: 9, the sequence of the downstream primer is SEQ ID NO: all primers were synthesized by Biotechnology engineering (Shanghai) Inc. The method comprises the following steps of: 6 as a template, carrying out PCR amplification to obtain a migratory locust V-ATPase-B gene fragment, purifying and cloning the obtained product, transforming the purified product into escherichia coli, and sending the escherichia coli to a company Limited in Biotechnology engineering (Shanghai) for sequencing, wherein the sequence of the migratory locust V-ATPase-B gene fragment is SEQ ID NO: 8.
according to SEQ ID NO: 11, designing a specific primer by using primer premier 5.0 software, wherein the sequence of an upstream primer is SEQ ID NO: 14, the sequence of the downstream primer is SEQ ID NO: all primers were synthesized by Biotechnology engineering (Shanghai) Inc. The method comprises the following steps of: 11 as a template, carrying out PCR amplification to obtain a migratory locust V-ATPase-C gene fragment, purifying and cloning the obtained product, transforming the purified product into escherichia coli, and sending the escherichia coli to a company Limited in Biotechnology engineering (Shanghai) for sequencing, wherein the sequence of the migratory locust V-ATPase-C gene fragment is SEQ ID NO: 13.
according to SEQ ID NO: 16, and designing a specific primer by using primer premier 5.0 software, wherein the sequence of an upstream primer is SEQ ID NO: 19, the sequence of the downstream primer is SEQ ID NO: all primers were synthesized by Biotechnology engineering (Shanghai) Inc. The method comprises the following steps of: 16 as a template, carrying out PCR amplification to obtain a migratory locust V-ATPase-A gene fragment, purifying and cloning the obtained product, transforming the purified product into escherichia coli, and sending the escherichia coli to a company Limited in Biotechnology engineering (Shanghai) for sequencing, wherein the sequence of the escherichia coli is SEQ ID NO: 18.
according to SEQ ID NO: 21, designing a specific primer by using primer premier 5.0 software, wherein the sequence of an upstream primer is SEQ ID NO: 24, the sequence of the downstream primer is SEQ ID NO: 25, all primers were synthesized by Biotechnology engineering (Shanghai) Inc. The method comprises the following steps of: 21 as a template, carrying out PCR amplification to obtain a migratory locust V-ATPase-A gene fragment, purifying and cloning the obtained product, transforming the purified product into escherichia coli, and sending the escherichia coli to a company Limited in Biotechnology engineering (Shanghai) for sequencing, wherein the sequence of the migratory locust V-ATPase-A gene fragment is SEQ ID NO: 23.
according to SEQ ID NO: 26, designing a specific primer by using primer premier 5.0 software, wherein the sequence of an upstream primer is SEQ ID NO: 29, the sequence of the downstream primer is SEQ ID NO: 30, all primers were synthesized by Biotechnology engineering (Shanghai) Inc. The method comprises the following steps of: 26 as a template, carrying out PCR amplification to obtain a migratory locust V-ATPase-A gene fragment, purifying and cloning the obtained product, transforming the purified product into escherichia coli, and sending the escherichia coli to a company Limited in Biotechnology engineering (Shanghai) for sequencing, wherein the sequence of the migratory locust V-ATPase-A gene fragment is SEQ ID NO: 28.
according to SEQ ID NO: 31, and designing a specific primer by using primer premier 5.0 software, wherein the sequence of an upstream primer is SEQ ID NO: 34, the sequence of the downstream primer is SEQ ID NO: 35, all primers were synthesized by Biotechnology engineering (Shanghai) Inc. The method comprises the following steps of: 31 as a template, carrying out PCR amplification to obtain a migratory locust V-ATPase-A gene fragment, purifying and cloning the obtained product, transforming the purified product into escherichia coli, and sending the escherichia coli to a company Limited in Biotechnology engineering (Shanghai) for sequencing, wherein the sequence of the migratory locust V-ATPase-A gene fragment is SEQ ID NO: 33.
example 2: synthesis of dsRNA (double-stranded ribonucleic acid) specific to each gene in V-ATPase-V1 domain of migratory locust
The sequences were determined using the sequences of SEQ ID NOs: 3. SEQ ID NO: 8. SEQ ID NO: 13. SEQ ID NO: 18. SEQ ID NO: 23. SEQ ID NO: 28. SEQ ID NO: 33 as a template, purified with Gel extraction kit (OMEGA) kit, and then purified according to HiScribeTMT7 High Yield RNA Synthesis Kit(NEWENGLAND
Figure BDA0002433412330000081
Inc.) kit to demonstrate in vitro transcription synthesis of the corresponding dsRNA quantitative determination was performed using NANODROP 2000(Thermo scientific) to a final concentration of 2. mu.g/. mu. L and stored in a super cryorefrigerator at-80 ℃ for future use.
Example 3: experiments on lethal locusts migratoria through dsRNA of each gene in V-ATPase-V1 domain
1) Injection of dsRNA of each gene of V-ATPase-V1 structural domain of migratory locust
L mV-ATPase-A gene dsRNA (ds L mV-ATPase-A) is injected into the 1 st day nymphs which are healthy in growth and consistent in size and are 72 years old, 3 mu L (6 mu g ds L mV-ATPase-A) is lightly injected into the space between two abdominal nodes and three abdominal nodes of the lateral abdomen of the nymph by using a micro injector, the same amount of dsGFP is injected into a control group at the same time, the locusts which are injected with the dsRNA are placed into an artificial climate box for raising (illumination: dark time is 14h:10h, temperature is 30 +/-2 ℃, humidity is 60 percent), and fresh wheat seedlings and wheat bran are fed every day.
The injection method of V-ATPase-B gene dsRNA (ds L mV-ATPase-B) and the injection amount of migratory locust are the same as those of V-ATPase-A gene dsRNA.
The injection method of V-ATPase-C gene dsRNA (ds L mV-ATPase-C) and the injection amount of migratory locust are the same as those of V-ATPase-A gene dsRNA.
The injection method of V-ATPase-D gene dsRNA (ds L mV-ATPase-D) and the injection amount of migratory locust are the same as those of V-ATPase-A gene dsRNA.
The injection method of V-ATPase-E gene dsRNA (ds L mV-ATPase-E) and the injection amount of migratory locust are the same as those of V-ATPase-A gene dsRNA.
The injection method of V-ATPase-F gene dsRNA (ds L mV-ATPase-F) and the injection amount of migratory locust are the same as those of V-ATPase-A gene dsRNA.
The injection method of V-ATPase-G gene dsRNA (ds L mV-ATPase-G) and the injection amount of migratory locust are the same as those of V-ATPase-A gene dsRNA.
2) Detection of silencing efficiency of each gene in V-ATPase-V1 structural domain of migratory locust
Collecting control group injected with dsGFP and treatment group injected with ds L mV-ATPase-A, ds L mV-ATPase-B, ds L0 mV-ATPase-C, ds L1 mV-ATPase-D, ds L2 mV-ATPase-E, ds L3 mV-ATPase-F and ds L mV-ATPase-G respectively, performing total RNA extraction on the nymphs at the time point of 24h, wherein each gene dsRNA group in the control and injection V-ATPase-V1 domains is 3 biological repeats, each biological repeat is 3 head test insects, performing reverse transcription to form first strand cDNA, detecting each target gene (L mV-ATPase-A, L mV-ATPase-B, ds L mV-ATPase-C, L mV-ATPase-D, L mV-ATPase-E, L-ATPase-F and L mV-ATPase-G) and internal reference gene (L4 mV-ATPase-364 mV-ATPase-G) by adopting RT-qPCR method, and calculating the relative silencing reaction amount of the genesSystem and program reference
Figure BDA0002433412330000091
Results showed that injection of test insects L mV-ATPase-A group ds L mV-ATPase-A showed a significant decrease in gene expression as compared to the control group<0.001 (FIG. 1), L mV-ATPase-B Gene expression was significantly reduced in injected ds L mV-ATPase-B group test insects<0.01 (FIG. 3), L mV-ATPase-C gene expression was significantly reduced in group test insects with ds L mV-ATPase-C injection<0.001 (FIG. 5), L mV-ATPase-D gene expression was significantly reduced in group test insects with ds L mV-ATPase-D injection<0.001 (FIG. 7), L mV-ATPase-E gene expression was significantly reduced in group test insects with ds L mV-ATPase-E injection<0.001 (FIG. 9), L mV-ATPase-F gene expression was significantly reduced in group test insects with ds L mV-ATPase-F injection<Test insects L mV-ATPase-G gene expression was significantly reduced in 0.001 (FIG. 11), injected ds L mV-ATPase-G group<0.001 (fig. 13).
3) Phenotypic observation of 3-year-old nymphs after injection of V-ATPase-V1 domain gene dsRNA
After 3-year-old nymphs inject ds L mV-ATPase-A, the control group worms successfully molt to 4 years on day 5 and the development state of the worms after molting is good, compared with the control group, the locusts after injection of ds L mV-ATPase-A have the mortality rate of 79.7 percent and have the phenotype that 1 is the ridge line of the locusts at the age of 3 years of the locusts at the age of 4 years of the locusts at the treatment group and opens and bends and dies, 2 is the locusts at the age of 3 years of the locusts at the treatment group and dies 1-2 days after the time of 4 years of the locusts at the age of 4, 3 is the locusts at the treatment group and dies 1-2 days before the time of 5 years of the locusts at the age of 4 of the locusts at the treatment group, and the.
After 3-year-old nymphs inject ds L mV-ATPase-B, the control group worms all successfully molt to 4 years on day 5, and the development state of the worms after molting is good, compared with the control group, the locusts migratorius after injection of ds L mV-ATPase-B have the mortality rate of 85.7 percent and have the phenotype of 1 death in 1-2 days before 3-year-old molting of the locusts migratorius of the treatment group at 4 years, 2 death in 3-year-old molting of the locusts migratorius of the treatment group at 4 years, 3 death in 1-2 days after 3-year old molting of the locusts of the treatment group at 4 years, and the results are shown in FIG. 4.
After 3-year-old nymphs inject ds L mV-ATPase-C, the control group worms all successfully molt to 4-year-old on day 5, and the development state of the worms after molting is good, compared with the control group, the locusts after injection of ds L mV-ATPase-C have the mortality rate of 69.8 percent, and have the phenotype that 1 is the treated locusts cannot successfully molt and die due to ridge line cracking when being molted for 4-year-old at 3-year-old, and 2 is the treated locusts die 1-2 days after being molted for 4-year old at 3-year old, and the result is shown in FIG. 6.
After 3-year-old nymphs inject ds L mV-ATPase-D, the control group worms successfully molt to 4 years on day 5, and the development state of the worms after molting is good, compared with the control group, the locusts migratorius after injection of ds L mV-ATPase-D have the mortality rate of 90.4 percent and have the phenotype that 1 is the death of the locusts migratorius in the treatment group 1-2 days before the 3-year-old molting of the locusts migratorius in the treatment group 4 years, 2 is the death of the locusts migratorius in the treatment group 3 years after the 4-year molting of the locusts in the treatment group cannot successfully slough off the old cuticle, and 3 is the death of the locusts migratorius in the treatment.
After 3-year-old nymphs inject ds L mV-ATPase-E, the control group worms all successfully molt to 4-year-old on day 5, and the development state of the worms after molting is good, compared with the control group, the locusts migratorius after injection of ds L mV-ATPase-E have the mortality rate of 93.5 percent and the phenotype of 1 death 1-2 days before 3-year-old molting of the locusts migratorius of the treatment group and 2 ridge line rupture and back bowing and death when 3-year-old molting of the locusts of the treatment group is 4-year old, and the results are shown in FIG. 10.
After 3-year-old nymphs inject ds L mV-ATPase-F, the control group worms successfully molt to 4 years on day 5 and the development state of the worms after molting is good, compared with the control group, the locusts after injection of ds L mV-ATPase-F have the mortality rate of 77.8 percent and have the following phenotype that 1 treated locusts cannot successfully slough off old epidermis when being aged 3 to 4 and 2 treated locusts die after being aged 4 and the result is shown in figure 12.
Compared with the control group, the locusts in the control group after injection of the ds L mV-ATPase-G have the mortality rate of 60.3 percent and have the phenotype that 1 is the death of the locusts in the treatment group 1-2 days before the 3 rd locusts in the period of 4 th, and 2 is the death of the old outer skins which cannot be successfully sloughed off when the locusts in the treatment group are at the 3 rd stage and the 4 th stage in the period of 4 th, and the result is shown in FIG. 14.
In conclusion, the V-ATPase-V1 structural domain gene fragments of the migratory locust comprise the gene fragments of V-ATPase-A, V-ATPase-B, V-ATPase-C, V-ATPase-D, V-ATPase-E, V-ATPase-F and V-ATPase-G, and the dsRNA synthesized by each gene fragment can silence target genes specifically after being injected into the body cavity of the migratory locust, so that the migratory locust is prevented from growing and developing and causes death, and the death rate reaches 60.3% -93.7%. Therefore, the dsRNA of the V-ATPase-V1 structural domain gene of the migratory locust is applied to the control of the migratory locust, and the specific implementation method is as follows: the dsRNA is prepared into a spraying pesticide, or the dsRNA is prepared into a bait, or the dsRNA is transferred into a migratory locust feeding plant body. Optionally, one or more dsRNA is/are prepared into a spraying pesticide or a bait, or the dsRNA is transferred into a feeding plant body of the migratory locust and is applied to the control of the migratory locust.
The above description is only for the specific embodiments of the present invention, but the scope of the present invention is not limited thereto, and any changes or substitutions that can be easily conceived by those skilled in the art within the technical scope of the present invention are included in the scope of the present invention. Therefore, the protection scope of the present invention shall be subject to the protection scope of the appended claims.
LmV-ATPase-A SEQ ID NO：1
atgacgagcacattgataaagacgtccgatgaggaccgggagtccaaattcggctttgtttttgccgtatctggacctgtggtgacagctgaacgaatggccggttctgctatgtacgaactggtgcgtgtcggttattatgaactggtcggagagatcatccggttggagggtgacatggcaacaatccaagtatacgaagacacctcaggtgtgacagtaggcgatcccgtgctgcgcacaggcaagccgctgtccgtggaactgggacccggaatcatgggcagcatcttcgacggtatccagcgaccgctgaaggatatcaatgaactgtcaaatagtatctacatcccgaaaggtgtcaatgtgcctgccctgagtcgcactgcacagtgggacttcagtcccgtcagtgtcaaggttggaagccacattactggtggtgacctgtacggtttggtccacgaaaatactctggtgaaacacaagttgctgctgccgccccgtgccaagggaactgtcacgtacattgcagaacctggaaactacacagttgatgatgttgtcctggagacagaatttgacggcgagcgatcaaagttcaccatgctgcaagtgtggcctgtacgtcagcccaggcctgttacagaaaagttgccagctaactaccccctccttactggccagcgtgtgctcgactccctattcccgtgtgtccagggtggaacaacagctattcctggggccttcggatgtggcaagactgtaatatcacagtctttgtcaaaatactcaaactccgatgtaattatctatgtaggttgtggtgagcgaggtaatgaaatgtcagaagtactcagggatttccccgagttgtcggtggagattgatggtgtgactgaatcaatcatgaagagaacagccctggtcgcaaacacatcaaacatgcctgtggctgctcgagaagcatctatctacacaggtattacactgtcagaatacttcagggacatgggttacaatgtatccatgatggctgactcaacttcacgatgggccgaagctcttcgagaaatctcaggtcgattggctgaaatgcctgccgacagcggttatcccgcctacctaggtgcacgacttgccagtttctacgagcgtgccggccgtgtgaagtgcttgggtaacccagacagggagggctccgtgagtatagtgggcgccgtgtcgccgcccggtggagacttctcagatcccgtgacgacggccacactaggtatcgtccaggtgttctggggtctcgacaagaaacttgcccagcgaaagcacttcccatccatcaactggctcatctcgtacagtaaatacatgcgtgctctggatgacttctacgacaagaatttcccagagtttgtcccactgcgtacaaaggtgaaggagattttgcaggaggaagaagacctgtctgaaattgtgcagttggtcggtaaagcttcattggcagaaactgacaagatcacacttgaggttgccaaactattaaaggatgatttcctgcaacagaacagctattcaccatatgaccgtttctgcccattctacaagacagtaggaatgctgaaaaatatgattgctttctacgatatgtctcggcatgcagttgaatctactgctcagagcgagaacaagatcacttggaatgttattagagattctatgggcaatattctgtatcagctttcctccatgaaattcaaggatccagtcaaggatggagaagcgaagatcaaggcagactttgagcagcttcatgaagacattcagcaagccttcaggaacctggaggattaa
LmV-ATPase-A SEQ ID NO：2
MTSTLIKTSDEDRESKFGFVFAVSGPVVTAERMAGSAMYELVRVGYYELVGEIIRLEGDMATIQVYEDTSGVTVGDPVLRTGKPLSVELGPGIMGSIFDGIQRPLKDINELSNSIYIPKGVNVPALSRTAQWDFSPVSVKVGSHITGGDLYGLVHENTLVKHKLLLPPRAKGTVTYIAEPGNYTVDDVVLETEFDGERSKFTMLQVWPVRQPRPVTEKLPANYPLLTGQRVLDSLFPCVQGGTTAIPGAFGCGKTVISQSLSKYSNSDVIIYVGCGERGNEMSEVLRDFPELSVEIDGVTESIMKRTALVANTSNMPVAAREASIYTGITLSEYFRDMGYNVSMMADSTSRWAEALREISGRLAEMPADSGYPAYLGARLASFYERAGRVKCLGNPDREGSVSIVGAVSPPGGDFSDPVTTATLGIVQVFWGLDKKLAQRKHFPSINWLISYSKYMRALDDFYDKNFPEFVPLRTKVKEILQEEEDLSEIVQLVGKASLAETDKITLEVAKLLKDDFLQQNSYSPYDRFCPFYKTVGMLKNMIAFYDMSRHAVESTAQSENKITWNVIRDSMGNILYQLSSMKFKDPVKDGEAKIKADFEQLHEDIQQAFRNLED
LmV-ATPase-A SEQ ID NO：3
gcaaacacatcaaacatgcctgtggctgctcgagaagcatctatctacacaggtattacactgtcagaatacttcagggacatgggttacaatgtatccatgatggctgactcaacttcacgatgggccgaagctcttcgagaaatctcaggtcgattggctgaaatgcctgccgacagcggttatcccgcctacctaggtgcacgacttgccagtttctacgagcgtgccggccgtgtgaagtgcttgggtaacccagacagggagggctccgtgagtatagtgggcgccgtgtcgccgcccggtggagacttctcagatcccgtgacgacggccacactaggtatcgtccaggtgttctggggtctcgacaagaaacttgcccagcgaaagcacttcccatccatcaactggctcatctcgtacagtaaatacatgcgtgctctggatgacttctacgacaagaatttcccagagtttgtcccactgcgtacaaaggt
LmV-ATPase-A SEQ ID NO：4
taatacgactcactataggggcaaacacatcaaacatgcc
LmV-ATPase-A SEQ ID NO：5
taatacgactcactatagggacctttgtacgcagtgggac
LmV-ATPase-B SEQ ID NO：6
atgtcgtacaacaaagttataagtccaaatcaggcgaataaagaacatgttttggcagtttcaagagattttatatcccagcctcgtctaacatataaaactgtatctggtgtcaatggaccactagttattctcgacgaagttaagttcccaaagttcgctgaaattgttcagttgaaacttgcagatggaactatcagatcgggccaggtcttggaagtcagtggatctaaagctgttgtccaagtttttgaaggtacatcaggtatagacgccaaaaatacactatgtgaattcacaggagatattctgcggacaccagtgtcagaggatatgttgggcagagtattcaatgggagtgggaaacccattgacaaaggcccacctattttggctgaagattatcttgatattcagggtcaacctattaacccatggtctcgtatatatccggaagaaatgatccagacaggtatatctgccattgatgtgatgaattccattgcccgtgggcaaaagattccaatcttctctgctgctggcttgccacacaatgaaattgctgctcagatctgccgtcaagcaggacttgttaagcttccaggaaagtcagttttggatgatcatgaggataattttgctatcgtatttgcagctatgggtgttaacatggagacagctcgtttcttcaagcaggactttgaggaaaatggttctatggaaaacgtgtgtctgttcttgaacttggccaatgatccaaccattgaacgtatcatcacaccccggcttgcattgacagctgctgaattcctggcatatcaatgtgagaaacacgtattagtcattctcacagacatgagttcatatgctgaggctttgcgagaggtttcagctgctagggaggaagtaccagggcgaagaggattccctggttacatgtacacagatcttgccaccatctatgaaagagctgggagagtggaaggtcgaagtggttccattacacagattcccattctgactatgcccaatgatgacattacacatcctatcccagatctgacaggatacatcacagaaggtcaaatatatgtagaccgtcagttgcacaacaggcaaatatatcctccagtcaatgtattgccatcactcagtcgtctcatgaagtcggccattggtgaaaacatgactcgtaaggaccatgctgatgtatcaaaccagctgtacgcttgttacgctattgggaaagatgtgcaggccatgaaggctgttgttggtgaggaagctcttaccccagatgaccttctgtatctcgaattcctcacaaaatttgagaagaatttcatatctcaaggtaactatgagaaccgtaccgtatttgaatcattggatattggttggcaactgcttcgaattttcccgaaggagatgttgaagaggatcccagcatcaatacttgcagaattctatccacgtgactcaagacaccctcaaactaaatag
LmV-ATPase-B SEQ ID NO：7
MSYNKVISPNQANKEHVLAVSRDFISQPRLTYKTVSGVNGPLVILDEVKFPKFAEIVQLKLADGTIRSGQVLEVSGSKAVVQVFEGTSGIDAKNTLCEFTGDILRTPVSEDMLGRVFNGSGKPIDKGPPILAEDYLDIQGQPINPWSRIYPEEMIQTGISAIDVMNSIARGQKIPIFSAAGLPHNEIAAQICRQAGLVKLPGKSVLDDHEDNFAIVFAAMGVNMETARFFKQDFEENGSMENVCLFLNLANDPTIERIITPRLALTAAEFLAYQCEKHVLVILTDMSSYAEALREVSAAREEVPGRRGFPGYMYTDLATIYERAGRVEGRSGSITQIPILTMPNDDITHPIPDLTGYITEGQIYVDRQLHNRQIYPPVNVLPSLSRLMKSAIGENMTRKDHADVSNQLYACYAIGKDVQAMKAVVGEEALTPDDLLYLEFLTKFEKNFISQGNYENRTVFESLDIGWQLLRIFPKEMLKRIPASILAEFYPRDSRHPQTK
LmV-ATPase-B SEQ ID NO：8
tgacagctgctgaattcctggcatatcaatgtgagaaacacgtattagtcattctcacagacatgagttcatatgctgaggctttgcgagaggtttcagctgctagggaggaagtaccagggcgaagaggattccctggttacatgtacacagatcttgccaccatctatgaaagagctgggagagtggaaggtcgaagtggttccattacacagattcccattctgactatgcccaatgatgacattacacatcctatcccagatctgacaggatacatcacagaaggtcaaatatatgtagaccgtcagttgcacaacaggcaaatatatcctccagtcaatgtattgccatcactcagtcgtctcatgaagtcggccattggtgaaaacatgactcgtaaggaccatgctgatgtatcaaaccagctgtacgcttgttacgctattgggaaagatgtgcaggccatgaaggctgttgttggtgag
LmV-ATPase-B SEQ ID NO：9
taatacgactcactatagggtgacagctgctgaattcctg
LmV-ATPase-B SEQ ID NO：10
taatacgactcactatagggctcaccaacaacagccttca
LmV-ATPase-C SEQ ID NO：11
atgacggaatactggctaatatcggctccaggagacaaaacttgccagcaaacatgggaaactttgaataatctgacgagtaaacagaacaatgtatcagtaaactacaaattccatattccagacctaaaggttggtacactggatcagctggttggtttatcagatgatttagggaagctagacagctatgttgaacaagtgaccaggaaggttgcagcctatcttggtgaagtactcgaagaccaaagagacaaactccatgaaaaccttttggcaaacaatagtgatcttcctgggtacatcacacgttttcagtgggatatggcaaaatatcccatcaaacaatctttgcggaacatagctgacattatcagtaaacaagtaggtcagatagatgctgatttaaaaacaaaatctactgcttataataatttgaaaggcaacctgcaaaatctggagaagaagcaaactgggagtctactaacgagaaatctcgctgatttagtgaagaaagagcacttcatattggattctgaataccttactacactattggttattgtaccaaaaatactattcaatgaatggaatcagaactatgagaaaattacagatatgattgtacctcgatcaagccaattgatttaccaggattcggactttggcttgtacactgtgactttgttcaagaaagttgttgatgaatttaaactccatgctcgagagagaaagtttgttgtaagagatttcacttacaatgaggaagaattgttggctgggaaaaatgaaatgacaaaactggttacggacaagaagaaacaatttggcccattagtacgctggctcaaagtcaacttcagtgaatgtttctgtgcatggatacatgttaaagcactgagagtgtttgtggaatctgttctgaggtatggactgcccgtgaacttccaagctatgctgcttcacccacacaaaaaaagtacgaagaggttacgagacctcctgaaccaattatatggtcatctagacagcagtgcctcacaagggctgggtgcacatgatagtgtggagataccaggacttgggtttggccatgctgagtattacccatatgtttattacaaaattaatgttgatatggttgatactaaagtgtaa
LmV-ATPase-C SEQ ID NO：12
MTEYWLISAPGDKTCQQTWETLNNLTSKQNNVSVNYKFHIPDLKVGTLDQLVGLSDDLGKLDSYVEQVTRKVAAYLGEVLEDQRDKLHENLLANNSDLPGYITRFQWDMAKYPIKQSLRNIADIISKQVGQIDADLKTKSTAYNNLKGNLQNLEKKQTGSLLTRNLADLVKKEHFILDSEYLTTLLVIVPKILFNEWNQNYEKITDMIVPRSSQLIYQDSDFGLYTVTLFKKVVDEFKLHARERKFVVRDFTYNEEELLAGKNEMTKLVTDKKKQFGPLVRWLKVNFSECFCAWIHVKALRVFVESVLRYGLPVNFQAMLLHPHKKSTKRLRDLLNQLYGHLDSSASQGLGAHDSVEIPGLGFGHAEYYPYVYYKINVDMVDTKV
LmV-ATPase-C SEQ ID NO：13
aaggcaacctgcaaaatctggagaagaagcaaactgggagtctactaacgagaaatctcgctgatttagtgaagaaagagcacttcatattggattctgaataccttactacactattggttattgtaccaaaaatactattcaatgaatggaatcagaactatgagaaaattacagatatgattgtacctcgatcaagccaattgatttaccaggattcggactttggcttgtacactgtgactttgttcaagaaagttgttgatgaatttaaactccatgctcgagagagaaagtttgttgtaagagatttcacttacaatgaggaagaattgttggctgggaaaaatgaaatgacaaaactggttacggacaagaagaaacaatttggcccattagtacgctggctcaaagtcaacttcagtgaatgtttctgtgcatggatacatgttaaagcactgagagtgtttgtggaatctgttctgaggtatggactgccc
LmV-ATPase-C SEQ ID NO：14
taatacgactcactatagggaaggcaacctgcaaaatctg
LmV-ATPase-C SEQ ID NO：15
taatacgactcactataggggggcagtccatacctcagaa
LmV-ATPase-D SEQ ID NO：16
atgtcgggaaaagacagaattcaaatatttccctcgcgaggtgcgcagatgttaatgaaagctagactaaaaggagcacaaaaaggtcacagtttactgaagaaaaaggcagatgctttgcaaatgcgatttcggcagatactgggcaaaatcatagagactaagacgctcatgggagaagtaatgaaagaagctgctttctcattagctgaagcaaaatttactaccggagatttcaaccaagttgttctgcagaacgtaaatagggcacaagtgaagattcgcacaaagaaggacaatgttgcaggtgttatgttaccagtatttgaaagttatcaagatggttcagacacatatgaactggcaggtttggcaagaggaggacagcagcttgccaaactaaaaaagaactaccagagtgcagtaaagttgttagttgaattagcttctctacagacatcttttgttactcttgacgaggtcatcaaaattaccaatagaagagttaatgcaattgaacatgtaattattccacgcattgaacgtactcttgcctacattatatctgaattggatgaattggagagagaagagttttaccggttgaagaaaattcaagacaagaaacgcatagccagagccaaagccgaggcgaaagcagaagcccagaagcaggccctcctgaaggcgggccaagacgtcacctacaccccgagcctgctcgacgaaggggacgaagacatcttgttctag
LmV-ATPase-D SEQ ID NO：17
MSGKDRIQIFPSRGAQMLMKARLKGAQKGHSLLKKKADALQMRFRQILGKIIETKTLMGEVMKEAAFSLAEAKFTTGDFNQVVLQNVNRAQVKIRTKKDNVAGVMLPVFESYQDGSDTYELAGLARGGQQLAKLKKNYQSAVKLLVELASLQTSFVTLDEVIKITNRRVNAIEHVIIPRIERTLAYIISELDELEREEFYRLKKIQDKKRIARAKAEAKAEAQKQALLKAGQDVTYTPSLLDEGDEDILF
LmV-ATPase-D SEQ ID NO：18
gcgcagatgttaatgaaagctagactaaaaggagcacaaaaaggtcacagtttactgaagaaaaaggcagatgctttgcaaatgcgatttcggcagatactgggcaaaatcatagagactaagacgctcatgggagaagtaatgaaagaagctgctttctcattagctgaagcaaaatttactaccggagatttcaaccaagttgttctgcagaacgtaaatagggcacaagtgaagattcgcacaaagaaggacaatgttgcaggtgttatgttaccagtatttgaaagttatcaagatggttcagacacatatgaactggcaggtttggcaagaggaggacagcagcttgccaaactaaaaaagaactaccagagtgcagtaaagttgttagttgaattagcttctctacagacatcttttgttactcttgacgaggtcatcaaaattaccaatagaagagttaatgcaattgaacatgtaattattccacgcattgaacgtactcttgcctac
LmV-ATPase-D SEQ ID NO：19
taatacgactcactataggggcgcagatgttaatgaaagc
LmV-ATPase-D SEQ ID NO：20
taatacgactcactataggggtaggcaagagtacgttca
LmV-ATPase-E SEQ ID NO：21
atggctctaagcgatgctgatgtccaaaaacagattaagcatatgatggcctttatagatcaagaggcaaatgaaaaggccgaagaaatagatgcaaaagctgaagaagaattcaatattgagaaaggccgattggtgcaacagcaacgtttaaaaataatggaatattatgaaagaaaggaaaaacaagtggaactccagaagaaaattcagtcttcaaacatgctaaaccaagcacgcttaaaagctctgaaagttcgtgaggatcatgtgcgtggtgtacttgaagaggcaagaaagagattgggggagatcacaaaggatcagtctcgatacagccatgttctccatcagttaataatgcaggggctgctgcagctcttggagccgaatgtcactgtaaggacacgtgaagttgaccagcgcattgttgatagcatcttgccagctattacacaaaagtacaaagaaattactggtgggaaggatatctcgctaaaagtggatactgaagccttcttgaacccagaagttactggtggaatagagttgctagcacaaaagggcagaattaaaattgttaatactttggaagcacggctggagctcattgcccagcaacttattcctgagatacgatgtgcattgtttggtcgtaacccaaatcgcaagttcgctgactaa
LmV-ATPase-E SEQ ID NO：22
MALSDADVQKQIKHMMAFIDQEANEKAEEIDAKAEEEFNIEKGRLVQQQRLKIMEYYERKEKQVELQKKIQSSNMLNQARLKALKVREDHVRGVLEEARKRLGEITKDQSRYSHVLHQLIMQGLLQLLEPNVTVRTREVDQRIVDSILPAITQKYKEITGGKDISLKVDTEAFLNPEVTGGIELLAQKGRIKIVNTLEARLELIAQQLIPEIRCALFGRNPNRKFAD
LmV-ATPase-E SEQ ID NO：23
ctaagcgatgctgatgtccaaaaacagattaagcatatgatggcctttatagatcaagaggcaaatgaaaaggccgaagaaatagatgcaaaagctgaagaagaattcaatattgagaaaggccgattggtgcaacagcaacgtttaaaaataatggaatattatgaaagaaaggaaaaacaagtggaactccagaagaaaattcagtcttcaaacatgctaaaccaagcacgcttaaaagctctgaaagttcgtgaggatcatgtgcgtggtgtacttgaagaggcaagaaagagattgggggagatcacaaaggatcagtctcgatacagccatgttctccatcagttaataatgcaggggctgctgcagctcttggagccgaatgtcactgta
LmV-ATPase-E SEQ ID NO：24
taatacgactcactatagggctaagcgatgctgatgtcca
LmV-ATPase-E SEQ ID NO：25
taatacgactcactatagggtacagtgacattcggctcca
LmV-ATPase-F SEQ ID NO：26
atggcattacattcggcttacaaagggaaactcgtctctgtcatcggagatgaggacacctgcgttggctttcttcttggaggagttggagaagtcaataaaagcagacatcccaatttcatggtggtggacaaaaacaccagtgttggtgagatagaagagtgcttcaaaagatttgtgaagagagatgatattgacatcatactgattaaccagaatattgctgagatgattcgacacgtaatcgacagccactcgcagcccgttccgtctgtcctcgagataccgtcaaaggaccatccttatgatgccaccaaagattcgatattacgtagggcaaagggaatgttcaacccagatgacttcaagtaa
LmV-ATPase-F SEQ ID NO：27
MALHSAYKGKLVSVIGDEDTCVGFLLGGVGEVNKSRHPNFMVVDKNTSVGEIEECFKRFVKRDDIDIILINQNIAEMIRHVIDSHSQPVPSVLEIPSKDHPYDATKDSILRRAKGMFNPDDFK
LmV-ATPase-F SEQ ID NO：28
acattcggcttacaaagggaaactcgtctctgtcatcggagatgaggacacctgcgttggctttcttcttggaggagttggagaagtcaataaaagcagacatcccaatttcatggtggtggacaaaaacaccagtgttggtgagatagaagagtgcttcaaaagatttgtgaagagagatgatattgacatcatactgattaaccagaatattgctgagatgattcgacacgtaatcgacagccactcgcagcccgttccgtctgtcctcgagataccgtcaaaggaccatcctta
LmV-ATPase-F SEQ ID NO：29
taatacgactcactatagggacattcggcttacaaaggga
LmV-ATPase-F SEQ ID NO：30
taatacgactcactatagggtaaggatggtcctttgacgg
LmV-ATPase-G SEQ ID NO：31
atggctagccagactcagggaattcaacagcttctagcagctgaaaaaagggcagcagagaaggtttcagaagcaagaaagcgaaaagcacgcaggttgaaacaggccaaagaagaagcccaggaagaaattgaaaaatacagacaagaaagagagaggcagtttaaagagtttgaagcaaagcacatgggctcacgagaggatagggctgcacagattgaagcagaaacaaaagtaaagattgcagaaatggagaaatctgtaaatcagcggaaggaaaagttgatagaaaggattttggaattggtgtacgatatcaagcccgaactccataagaacttcagagctgcaaagtag
LmV-ATPase-G SEQ ID NO：32
MASQTQGIQQLLAAEKRAAEKVSEARKRKARRLKQAKEEAQEEIEKYRQERERQFKEFEAKHMGSREDRAAQIEAETKVKIAEMEKSVNQRKEKLIERILELVYDIKPELHKNFRAAK
LmV-ATPase-G SEQ ID NO：33
aagggcagcagagaaggtttcagaagcaagaaagcgaaaagcacgcaggttgaaacaggccaaagaagaagcccaggaagaaattgaaaaatacagacaagaaagagagaggcagtttaaagagtttgaagcaaagcacatgggctcacgagaggatagggctgcacagattgaagcagaaacaaaagtaaagattgcagaaatggagaaatctgtaaatcagcggaaggaaaagttgat
LmV-ATPase-G SEQ ID NO：34
taatacgactcactatagggaagggcagcagagaaggttt
LmV-ATPase-G SEQ ID NO：35
taatacgactcactatagggatcaacttttccttccgctg
Sequence listing
<110> university of Shanxi
V-ATPase-V1 structural domain gene of <120> migratory locust and application of dsRNA thereof in pest control
<160>35
<170>SIPOSequenceListing 1.0
<210>1
<211>1848
<212>DNA
<213> migratory locust (L austa migratoria)
<400>1
atgacgagca cattgataaa gacgtccgat gaggaccggg agtccaaatt cggctttgtt 60
tttgccgtat ctggacctgt ggtgacagct gaacgaatgg ccggttctgc tatgtacgaa 120
ctggtgcgtg tcggttatta tgaactggtc ggagagatca tccggttgga gggtgacatg 180
gcaacaatcc aagtatacga agacacctca ggtgtgacag taggcgatcc cgtgctgcgc 240
acaggcaagc cgctgtccgt ggaactggga cccggaatca tgggcagcat cttcgacggt 300
atccagcgac cgctgaagga tatcaatgaa ctgtcaaata gtatctacat cccgaaaggt 360
gtcaatgtgc ctgccctgag tcgcactgca cagtgggact tcagtcccgt cagtgtcaag 420
gttggaagcc acattactgg tggtgacctg tacggtttgg tccacgaaaa tactctggtg 480
aaacacaagt tgctgctgcc gccccgtgcc aagggaactg tcacgtacat tgcagaacct 540
ggaaactaca cagttgatga tgttgtcctg gagacagaat ttgacggcga gcgatcaaag 600
ttcaccatgc tgcaagtgtg gcctgtacgt cagcccaggc ctgttacaga aaagttgcca 660
gctaactacc ccctccttac tggccagcgt gtgctcgact ccctattccc gtgtgtccag 720
ggtggaacaa cagctattcc tggggccttc ggatgtggca agactgtaat atcacagtct 780
ttgtcaaaat actcaaactc cgatgtaatt atctatgtag gttgtggtga gcgaggtaat 840
gaaatgtcag aagtactcag ggatttcccc gagttgtcgg tggagattga tggtgtgact 900
gaatcaatca tgaagagaac agccctggtc gcaaacacat caaacatgcc tgtggctgct 960
cgagaagcat ctatctacac aggtattaca ctgtcagaat acttcaggga catgggttac 1020
aatgtatcca tgatggctga ctcaacttca cgatgggccg aagctcttcg agaaatctca 1080
ggtcgattgg ctgaaatgcc tgccgacagc ggttatcccg cctacctagg tgcacgactt 1140
gccagtttct acgagcgtgc cggccgtgtg aagtgcttgg gtaacccaga cagggagggc 1200
tccgtgagta tagtgggcgc cgtgtcgccg cccggtggag acttctcaga tcccgtgacg 1260
acggccacac taggtatcgt ccaggtgttc tggggtctcg acaagaaact tgcccagcga 1320
aagcacttcc catccatcaa ctggctcatc tcgtacagta aatacatgcg tgctctggat 1380
gacttctacg acaagaattt cccagagttt gtcccactgc gtacaaaggt gaaggagatt 1440
ttgcaggagg aagaagacct gtctgaaatt gtgcagttgg tcggtaaagc ttcattggca 1500
gaaactgaca agatcacact tgaggttgcc aaactattaa aggatgattt cctgcaacag 1560
aacagctatt caccatatga ccgtttctgc ccattctaca agacagtagg aatgctgaaa 1620
aatatgattg ctttctacga tatgtctcgg catgcagttg aatctactgc tcagagcgag 1680
aacaagatca cttggaatgt tattagagat tctatgggca atattctgta tcagctttcc 1740
tccatgaaat tcaaggatcc agtcaaggat ggagaagcga agatcaaggc agactttgag 1800
cagcttcatg aagacattca gcaagccttc aggaacctgg aggattaa 1848
<210>2
<211>615
<212>PRT
<213> migratory locust (L austa migratoria)
<400>2
Met Thr Ser Thr Leu Ile Lys Thr Ser Asp Glu Asp Arg Glu Ser Lys
1 5 10 15
Phe Gly Phe Val Phe Ala Val Ser Gly Pro Val Val Thr Ala Glu Arg
20 25 30
Met Ala Gly Ser Ala Met Tyr Glu Leu Val Arg Val Gly Tyr Tyr Glu
35 40 45
Leu Val Gly Glu Ile Ile Arg Leu Glu Gly Asp Met Ala Thr Ile Gln
50 55 60
Val Tyr Glu Asp Thr Ser Gly Val Thr Val Gly Asp Pro Val Leu Arg
65 70 75 80
Thr Gly Lys Pro Leu Ser Val Glu Leu Gly Pro Gly Ile Met Gly Ser
85 90 95
Ile Phe Asp Gly Ile Gln Arg Pro Leu Lys Asp Ile Asn Glu Leu Ser
100 105 110
Asn Ser Ile Tyr Ile Pro Lys Gly Val Asn Val Pro Ala Leu Ser Arg
115 120 125
Thr Ala Gln Trp Asp Phe Ser Pro Val Ser Val Lys Val Gly Ser His
130 135 140
Ile Thr Gly Gly Asp Leu Tyr Gly Leu Val His Glu Asn Thr Leu Val
145 150 155 160
Lys His Lys Leu Leu Leu Pro Pro Arg Ala Lys Gly Thr Val Thr Tyr
165 170 175
Ile Ala Glu Pro Gly Asn Tyr Thr Val Asp Asp Val Val Leu Glu Thr
180 185 190
Glu Phe Asp Gly Glu Arg Ser Lys Phe Thr Met Leu Gln Val Trp Pro
195 200 205
Val Arg Gln Pro Arg Pro Val Thr Glu Lys Leu Pro Ala Asn Tyr Pro
210 215 220
Leu Leu Thr Gly Gln Arg Val Leu Asp Ser Leu Phe Pro Cys Val Gln
225 230 235 240
Gly Gly Thr Thr Ala Ile Pro Gly Ala Phe Gly Cys Gly Lys Thr Val
245 250 255
Ile Ser Gln Ser Leu Ser Lys Tyr Ser Asn Ser Asp Val Ile Ile Tyr
260 265 270
Val Gly Cys Gly Glu Arg Gly Asn Glu Met Ser Glu Val Leu Arg Asp
275 280 285
Phe Pro Glu Leu Ser Val Glu Ile Asp Gly Val Thr Glu Ser Ile Met
290 295 300
Lys Arg Thr Ala Leu Val Ala Asn Thr Ser Asn Met Pro Val Ala Ala
305 310 315 320
Arg Glu Ala Ser Ile Tyr Thr Gly Ile Thr Leu Ser Glu Tyr Phe Arg
325 330 335
Asp Met Gly Tyr Asn Val Ser Met Met Ala Asp Ser Thr Ser Arg Trp
340 345 350
Ala Glu Ala Leu Arg Glu Ile Ser Gly Arg Leu Ala Glu Met Pro Ala
355 360 365
Asp Ser Gly Tyr Pro Ala Tyr Leu Gly Ala Arg Leu Ala Ser Phe Tyr
370 375 380
Glu Arg Ala Gly Arg Val Lys Cys Leu Gly Asn Pro Asp Arg Glu Gly
385 390 395 400
Ser Val Ser Ile Val Gly Ala Val Ser Pro Pro Gly Gly Asp Phe Ser
405 410 415
Asp Pro Val Thr Thr Ala Thr Leu Gly Ile Val Gln Val Phe Trp Gly
420 425 430
Leu Asp Lys Lys Leu Ala Gln Arg Lys His Phe Pro Ser Ile Asn Trp
435 440 445
Leu Ile Ser Tyr Ser Lys Tyr Met Arg Ala Leu Asp Asp Phe Tyr Asp
450 455 460
Lys Asn Phe Pro Glu Phe Val Pro Leu Arg Thr Lys Val Lys Glu Ile
465 470 475 480
Leu Gln Glu Glu Glu Asp Leu Ser Glu Ile Val Gln Leu Val Gly Lys
485 490 495
Ala Ser Leu Ala Glu Thr Asp Lys Ile Thr Leu Glu Val Ala Lys Leu
500 505 510
Leu Lys Asp Asp Phe Leu Gln Gln Asn Ser Tyr Ser Pro Tyr Asp Arg
515 520 525
Phe Cys Pro Phe Tyr Lys Thr Val Gly Met Leu Lys Asn Met Ile Ala
530 535 540
PheTyr Asp Met Ser Arg His Ala Val Glu Ser Thr Ala Gln Ser Glu
545 550 555 560
Asn Lys Ile Thr Trp Asn Val Ile Arg Asp Ser Met Gly Asn Ile Leu
565 570 575
Tyr Gln Leu Ser Ser Met Lys Phe Lys Asp Pro Val Lys Asp Gly Glu
580 585 590
Ala Lys Ile Lys Ala Asp Phe Glu Gln Leu His Glu Asp Ile Gln Gln
595 600 605
Ala Phe Arg Asn Leu Glu Asp
610 615
<210>3
<211>500
<212>DNA
<213> migratory locust (L austa migratoria)
<400>3
gcaaacacat caaacatgcc tgtggctgct cgagaagcat ctatctacac aggtattaca 60
ctgtcagaat acttcaggga catgggttac aatgtatcca tgatggctga ctcaacttca 120
cgatgggccg aagctcttcg agaaatctca ggtcgattgg ctgaaatgcc tgccgacagc 180
ggttatcccg cctacctagg tgcacgactt gccagtttct acgagcgtgc cggccgtgtg 240
aagtgcttgg gtaacccaga cagggagggc tccgtgagta tagtgggcgc cgtgtcgccg 300
cccggtggag acttctcaga tcccgtgacg acggccacac taggtatcgt ccaggtgttc 360
tggggtctcg acaagaaact tgcccagcga aagcacttcc catccatcaa ctggctcatc 420
tcgtacagta aatacatgcg tgctctggat gacttctacg acaagaattt cccagagttt 480
gtcccactgc gtacaaaggt 500
<210>4
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>4
taatacgact cactataggg gcaaacacat caaacatgcc 40
<210>5
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>5
taatacgact cactataggg acctttgtac gcagtgggac 40
<210>6
<211>1503
<212>DNA
<213> migratory locust (L austa migratoria)
<400>6
atgtcgtaca acaaagttat aagtccaaat caggcgaata aagaacatgt tttggcagtt 60
tcaagagatt ttatatccca gcctcgtcta acatataaaa ctgtatctgg tgtcaatgga 120
ccactagtta ttctcgacga agttaagttc ccaaagttcg ctgaaattgt tcagttgaaa 180
cttgcagatg gaactatcag atcgggccag gtcttggaag tcagtggatc taaagctgtt 240
gtccaagttt ttgaaggtac atcaggtata gacgccaaaa atacactatg tgaattcaca 300
ggagatattc tgcggacacc agtgtcagag gatatgttgg gcagagtatt caatgggagt 360
gggaaaccca ttgacaaagg cccacctatt ttggctgaag attatcttga tattcagggt 420
caacctatta acccatggtc tcgtatatat ccggaagaaa tgatccagac aggtatatct 480
gccattgatg tgatgaattc cattgcccgt gggcaaaaga ttccaatctt ctctgctgct 540
ggcttgccac acaatgaaat tgctgctcag atctgccgtc aagcaggact tgttaagctt 600
ccaggaaagt cagttttgga tgatcatgag gataattttg ctatcgtatt tgcagctatg 660
ggtgttaaca tggagacagc tcgtttcttc aagcaggact ttgaggaaaa tggttctatg 720
gaaaacgtgt gtctgttctt gaacttggcc aatgatccaa ccattgaacg tatcatcaca 780
ccccggcttg cattgacagc tgctgaattc ctggcatatc aatgtgagaa acacgtatta 840
gtcattctca cagacatgag ttcatatgct gaggctttgc gagaggtttc agctgctagg 900
gaggaagtac cagggcgaag aggattccct ggttacatgt acacagatct tgccaccatc 960
tatgaaagag ctgggagagt ggaaggtcga agtggttcca ttacacagat tcccattctg 1020
actatgccca atgatgacat tacacatcct atcccagatc tgacaggata catcacagaa 1080
ggtcaaatat atgtagaccg tcagttgcac aacaggcaaa tatatcctcc agtcaatgta 1140
ttgccatcac tcagtcgtct catgaagtcg gccattggtg aaaacatgac tcgtaaggac 1200
catgctgatg tatcaaacca gctgtacgct tgttacgcta ttgggaaaga tgtgcaggcc 1260
atgaaggctg ttgttggtga ggaagctctt accccagatg accttctgta tctcgaattc 1320
ctcacaaaat ttgagaagaa tttcatatct caaggtaact atgagaaccg taccgtattt 1380
gaatcattgg atattggttg gcaactgctt cgaattttcc cgaaggagat gttgaagagg 1440
atcccagcat caatacttgc agaattctat ccacgtgact caagacaccc tcaaactaaa 1500
tag 1503
<210>7
<211>500
<212>PRT
<213> migratory locust (L austa migratoria)
<400>7
Met Ser Tyr Asn Lys Val Ile Ser Pro Asn Gln Ala Asn Lys Glu His
1 5 10 15
Val Leu Ala Val Ser Arg Asp Phe Ile Ser Gln Pro Arg Leu Thr Tyr
20 25 30
Lys Thr Val Ser Gly Val Asn Gly Pro Leu Val Ile Leu Asp Glu Val
35 40 45
Lys Phe Pro Lys Phe Ala Glu Ile Val Gln Leu Lys Leu Ala Asp Gly
50 55 60
Thr Ile Arg Ser Gly Gln Val Leu Glu Val Ser Gly Ser Lys Ala Val
65 70 75 80
Val Gln Val Phe Glu Gly Thr Ser Gly Ile Asp Ala Lys Asn Thr Leu
85 90 95
Cys Glu Phe Thr Gly Asp Ile Leu Arg Thr Pro Val Ser Glu Asp Met
100 105110
Leu Gly Arg Val Phe Asn Gly Ser Gly Lys Pro Ile Asp Lys Gly Pro
115 120 125
Pro Ile Leu Ala Glu Asp Tyr Leu Asp Ile Gln Gly Gln Pro Ile Asn
130 135 140
Pro Trp Ser Arg Ile Tyr Pro Glu Glu Met Ile Gln Thr Gly Ile Ser
145 150 155 160
Ala Ile Asp Val Met Asn Ser Ile Ala Arg Gly Gln Lys Ile Pro Ile
165 170 175
Phe Ser Ala Ala Gly Leu Pro His Asn Glu Ile Ala Ala Gln Ile Cys
180 185 190
Arg Gln Ala Gly Leu Val Lys Leu Pro Gly Lys Ser Val Leu Asp Asp
195 200 205
His Glu Asp Asn Phe Ala Ile Val Phe Ala Ala Met Gly Val Asn Met
210 215 220
Glu Thr Ala Arg Phe Phe Lys Gln Asp Phe Glu Glu Asn Gly Ser Met
225 230 235 240
Glu Asn Val Cys Leu Phe Leu Asn Leu Ala Asn Asp Pro Thr Ile Glu
245 250 255
Arg Ile Ile Thr Pro Arg Leu Ala Leu Thr Ala Ala Glu Phe Leu Ala
260 265270
Tyr Gln Cys Glu Lys His Val Leu Val Ile Leu Thr Asp Met Ser Ser
275 280 285
Tyr Ala Glu Ala Leu Arg Glu Val Ser Ala Ala Arg Glu Glu Val Pro
290 295 300
Gly Arg Arg Gly Phe Pro Gly Tyr Met Tyr Thr Asp Leu Ala Thr Ile
305 310 315 320
Tyr Glu Arg Ala Gly Arg Val Glu Gly Arg Ser Gly Ser Ile Thr Gln
325 330 335
Ile Pro Ile Leu Thr Met Pro Asn Asp Asp Ile Thr His Pro Ile Pro
340 345 350
Asp Leu Thr Gly Tyr Ile Thr Glu Gly Gln Ile Tyr Val Asp Arg Gln
355 360 365
Leu His Asn Arg Gln Ile Tyr Pro Pro Val Asn Val Leu Pro Ser Leu
370 375 380
Ser Arg Leu Met Lys Ser Ala Ile Gly Glu Asn Met Thr Arg Lys Asp
385 390 395 400
His Ala Asp Val Ser Asn Gln Leu Tyr Ala Cys Tyr Ala Ile Gly Lys
405 410 415
Asp Val Gln Ala Met Lys Ala Val Val Gly Glu Glu Ala Leu Thr Pro
420 425 430
Asp Asp Leu Leu Tyr Leu Glu Phe Leu Thr Lys Phe Glu Lys Asn Phe
435 440 445
Ile Ser Gln Gly Asn Tyr Glu Asn Arg Thr Val Phe Glu Ser Leu Asp
450 455 460
Ile Gly Trp Gln Leu Leu Arg Ile Phe Pro Lys Glu Met Leu Lys Arg
465 470 475 480
Ile Pro Ala Ser Ile Leu Ala Glu Phe Tyr Pro Arg Asp Ser Arg His
485 490 495
Pro Gln Thr Lys
500
<210>8
<211>488
<212>DNA
<213> migratory locust (L austa migratoria)
<400>8
tgacagctgc tgaattcctg gcatatcaat gtgagaaaca cgtattagtc attctcacag 60
acatgagttc atatgctgag gctttgcgag aggtttcagc tgctagggag gaagtaccag 120
ggcgaagagg attccctggt tacatgtaca cagatcttgc caccatctat gaaagagctg 180
ggagagtgga aggtcgaagt ggttccatta cacagattcc cattctgact atgcccaatg 240
atgacattac acatcctatc ccagatctga caggatacat cacagaaggt caaatatatg 300
tagaccgtca gttgcacaac aggcaaatat atcctccagt caatgtattg ccatcactca 360
gtcgtctcat gaagtcggcc attggtgaaa acatgactcg taaggaccat gctgatgtat420
caaaccagct gtacgcttgt tacgctattg ggaaagatgt gcaggccatg aaggctgttg 480
ttggtgag 488
<210>9
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>9
taatacgact cactataggg tgacagctgc tgaattcctg 40
<210>10
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>10
taatacgact cactataggg ctcaccaaca acagccttca 40
<210>11
<211>1158
<212>DNA
<213> migratory locust (L austa migratoria)
<400>11
atgacggaat actggctaat atcggctcca ggagacaaaa cttgccagca aacatgggaa 60
actttgaata atctgacgag taaacagaac aatgtatcag taaactacaa attccatatt 120
ccagacctaa aggttggtac actggatcag ctggttggtt tatcagatga tttagggaag 180
ctagacagct atgttgaaca agtgaccagg aaggttgcag cctatcttgg tgaagtactc 240
gaagaccaaa gagacaaact ccatgaaaac cttttggcaa acaatagtga tcttcctggg 300
tacatcacacgttttcagtg ggatatggca aaatatccca tcaaacaatc tttgcggaac 360
atagctgaca ttatcagtaa acaagtaggt cagatagatg ctgatttaaa aacaaaatct 420
actgcttata ataatttgaa aggcaacctg caaaatctgg agaagaagca aactgggagt 480
ctactaacga gaaatctcgc tgatttagtg aagaaagagc acttcatatt ggattctgaa 540
taccttacta cactattggt tattgtacca aaaatactat tcaatgaatg gaatcagaac 600
tatgagaaaa ttacagatat gattgtacct cgatcaagcc aattgattta ccaggattcg 660
gactttggct tgtacactgt gactttgttc aagaaagttg ttgatgaatt taaactccat 720
gctcgagaga gaaagtttgt tgtaagagat ttcacttaca atgaggaaga attgttggct 780
gggaaaaatg aaatgacaaa actggttacg gacaagaaga aacaatttgg cccattagta 840
cgctggctca aagtcaactt cagtgaatgt ttctgtgcat ggatacatgt taaagcactg 900
agagtgtttg tggaatctgt tctgaggtat ggactgcccg tgaacttcca agctatgctg 960
cttcacccac acaaaaaaag tacgaagagg ttacgagacc tcctgaacca attatatggt 1020
catctagaca gcagtgcctc acaagggctg ggtgcacatg atagtgtgga gataccagga 1080
cttgggtttg gccatgctga gtattaccca tatgtttatt acaaaattaa tgttgatatg 1140
gttgatacta aagtgtaa 1158
<210>12
<211>385
<212>PRT
<213> migratory locust (L austa migratoria)
<400>12
Met Thr Glu Tyr Trp Leu Ile Ser Ala Pro Gly Asp Lys Thr Cys Gln
1 5 10 15
Gln Thr Trp Glu Thr Leu Asn Asn Leu Thr Ser Lys Gln Asn Asn Val
20 25 30
Ser Val Asn Tyr Lys Phe His Ile Pro Asp Leu Lys Val Gly Thr Leu
35 40 45
Asp Gln Leu Val Gly Leu Ser Asp Asp Leu Gly Lys Leu Asp Ser Tyr
50 55 60
Val Glu Gln Val Thr Arg Lys Val Ala Ala Tyr Leu Gly Glu Val Leu
65 70 75 80
Glu Asp Gln Arg Asp Lys Leu His Glu Asn Leu Leu Ala Asn Asn Ser
85 90 95
Asp Leu Pro Gly Tyr Ile Thr Arg Phe Gln Trp Asp Met Ala Lys Tyr
100 105 110
Pro Ile Lys Gln Ser Leu Arg Asn Ile Ala Asp Ile Ile Ser Lys Gln
115 120 125
Val Gly Gln Ile Asp Ala Asp Leu Lys Thr Lys Ser Thr Ala Tyr Asn
130 135 140
Asn Leu Lys Gly Asn Leu Gln Asn Leu Glu Lys Lys Gln Thr Gly Ser
145 150 155 160
Leu Leu Thr ArgAsn Leu Ala Asp Leu Val Lys Lys Glu His Phe Ile
165 170 175
Leu Asp Ser Glu Tyr Leu Thr Thr Leu Leu Val Ile Val Pro Lys Ile
180 185 190
Leu Phe Asn Glu Trp Asn Gln Asn Tyr Glu Lys Ile Thr Asp Met Ile
195 200 205
Val Pro Arg Ser Ser Gln Leu Ile Tyr Gln Asp Ser Asp Phe Gly Leu
210 215 220
Tyr Thr Val Thr Leu Phe Lys Lys Val Val Asp Glu Phe Lys Leu His
225 230 235 240
Ala Arg Glu Arg Lys Phe Val Val Arg Asp Phe Thr Tyr Asn Glu Glu
245 250 255
Glu Leu Leu Ala Gly Lys Asn Glu Met Thr Lys Leu Val Thr Asp Lys
260 265 270
Lys Lys Gln Phe Gly Pro Leu Val Arg Trp Leu Lys Val Asn Phe Ser
275 280 285
Glu Cys Phe Cys Ala Trp Ile His Val Lys Ala Leu Arg Val Phe Val
290 295 300
Glu Ser Val Leu Arg Tyr Gly Leu Pro Val Asn Phe Gln Ala Met Leu
305 310 315 320
Leu His Pro His Lys LysSer Thr Lys Arg Leu Arg Asp Leu Leu Asn
325 330 335
Gln Leu Tyr Gly His Leu Asp Ser Ser Ala Ser Gln Gly Leu Gly Ala
340 345 350
His Asp Ser Val Glu Ile Pro Gly Leu Gly Phe Gly His Ala Glu Tyr
355 360 365
Tyr Pro Tyr Val Tyr Tyr Lys Ile Asn Val Asp Met Val Asp Thr Lys
370 375 380
Val
385
<210>13
<211>500
<212>DNA
<213> migratory locust (L austa migratoria)
<400>13
aaggcaacct gcaaaatctg gagaagaagc aaactgggag tctactaacg agaaatctcg 60
ctgatttagt gaagaaagag cacttcatat tggattctga ataccttact acactattgg 120
ttattgtacc aaaaatacta ttcaatgaat ggaatcagaa ctatgagaaa attacagata 180
tgattgtacc tcgatcaagc caattgattt accaggattc ggactttggc ttgtacactg 240
tgactttgtt caagaaagtt gttgatgaat ttaaactcca tgctcgagag agaaagtttg 300
ttgtaagaga tttcacttac aatgaggaag aattgttggc tgggaaaaat gaaatgacaa 360
aactggttac ggacaagaag aaacaatttg gcccattagt acgctggctc aaagtcaact 420
tcagtgaatg tttctgtgca tggatacatg ttaaagcactgagagtgttt gtggaatctg 480
ttctgaggta tggactgccc 500
<210>14
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>14
taatacgact cactataggg aaggcaacct gcaaaatctg 40
<210>15
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>15
taatacgact cactataggg gggcagtcca tacctcagaa 40
<210>16
<211>753
<212>DNA
<213> migratory locust (L austa migratoria)
<400>16
atgtcgggaa aagacagaat tcaaatattt ccctcgcgag gtgcgcagat gttaatgaaa 60
gctagactaa aaggagcaca aaaaggtcac agtttactga agaaaaaggc agatgctttg 120
caaatgcgat ttcggcagat actgggcaaa atcatagaga ctaagacgct catgggagaa 180
gtaatgaaag aagctgcttt ctcattagct gaagcaaaat ttactaccgg agatttcaac 240
caagttgttc tgcagaacgt aaatagggca caagtgaaga ttcgcacaaa gaaggacaat 300
gttgcaggtg ttatgttacc agtatttgaa agttatcaag atggttcaga cacatatgaa 360
ctggcaggtt tggcaagagg aggacagcag cttgccaaac taaaaaagaa ctaccagagt 420
gcagtaaagt tgttagttga attagcttct ctacagacat cttttgttac tcttgacgag 480
gtcatcaaaa ttaccaatag aagagttaat gcaattgaac atgtaattat tccacgcatt 540
gaacgtactc ttgcctacat tatatctgaa ttggatgaat tggagagaga agagttttac 600
cggttgaaga aaattcaaga caagaaacgc atagccagag ccaaagccga ggcgaaagca 660
gaagcccaga agcaggccct cctgaaggcg ggccaagacg tcacctacac cccgagcctg 720
ctcgacgaag gggacgaaga catcttgttc tag 753
<210>17
<211>250
<212>PRT
<213> migratory locust (L austa migratoria)
<400>17
Met Ser Gly Lys Asp Arg Ile Gln Ile Phe Pro Ser Arg Gly Ala Gln
1 5 10 15
Met Leu Met Lys Ala Arg Leu Lys Gly Ala Gln Lys Gly His Ser Leu
20 25 30
Leu Lys Lys Lys Ala Asp Ala Leu Gln Met Arg Phe Arg Gln Ile Leu
35 40 45
Gly Lys Ile Ile Glu Thr Lys Thr Leu Met Gly Glu Val Met Lys Glu
50 55 60
Ala Ala Phe Ser Leu Ala Glu Ala Lys Phe Thr Thr Gly Asp Phe Asn
65 70 75 80
Gln Val Val Leu Gln Asn Val Asn Arg Ala Gln Val Lys Ile Arg Thr
85 90 95
Lys Lys Asp Asn Val Ala Gly Val Met Leu Pro Val Phe Glu Ser Tyr
100 105 110
Gln Asp Gly Ser Asp Thr Tyr Glu Leu Ala Gly Leu Ala Arg Gly Gly
115 120 125
Gln Gln Leu Ala Lys Leu Lys Lys Asn Tyr Gln Ser Ala Val Lys Leu
130 135 140
Leu Val Glu Leu Ala Ser Leu Gln Thr Ser Phe Val Thr Leu Asp Glu
145 150 155 160
Val Ile Lys Ile Thr Asn Arg Arg Val Asn Ala Ile Glu His Val Ile
165 170 175
Ile Pro Arg Ile Glu Arg Thr Leu Ala Tyr Ile Ile Ser Glu Leu Asp
180 185 190
Glu Leu Glu Arg Glu Glu Phe Tyr Arg Leu Lys Lys Ile Gln Asp Lys
195 200 205
Lys Arg Ile Ala Arg Ala Lys Ala Glu Ala Lys Ala Glu Ala Gln Lys
210 215 220
Gln Ala Leu Leu Lys Ala Gly Gln Asp Val Thr Tyr Thr Pro Ser Leu
225230 235 240
Leu Asp Glu Gly Asp Glu Asp Ile Leu Phe
245 250
<210>18
<211>516
<212>DNA
<213> migratory locust (L austa migratoria)
<400>18
gcgcagatgt taatgaaagc tagactaaaa ggagcacaaa aaggtcacag tttactgaag 60
aaaaaggcag atgctttgca aatgcgattt cggcagatac tgggcaaaat catagagact 120
aagacgctca tgggagaagt aatgaaagaa gctgctttct cattagctga agcaaaattt 180
actaccggag atttcaacca agttgttctg cagaacgtaa atagggcaca agtgaagatt 240
cgcacaaaga aggacaatgt tgcaggtgtt atgttaccag tatttgaaag ttatcaagat 300
ggttcagaca catatgaact ggcaggtttg gcaagaggag gacagcagct tgccaaacta 360
aaaaagaact accagagtgc agtaaagttg ttagttgaat tagcttctct acagacatct 420
tttgttactc ttgacgaggt catcaaaatt accaatagaa gagttaatgc aattgaacat 480
gtaattattc cacgcattga acgtactctt gcctac 516
<210>19
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>19
taatacgact cactataggg gcgcagatgt taatgaaagc 40
<210>20
<211>39
<212>DNA
<213> migratory locust (L austa migratoria)
<400>20
taatacgact cactataggg gtaggcaaga gtacgttca 39
<210>21
<211>684
<212>DNA
<213> migratory locust (L austa migratoria)
<400>21
atggctctaa gcgatgctga tgtccaaaaa cagattaagc atatgatggc ctttatagat 60
caagaggcaa atgaaaaggc cgaagaaata gatgcaaaag ctgaagaaga attcaatatt 120
gagaaaggcc gattggtgca acagcaacgt ttaaaaataa tggaatatta tgaaagaaag 180
gaaaaacaag tggaactcca gaagaaaatt cagtcttcaa acatgctaaa ccaagcacgc 240
ttaaaagctc tgaaagttcg tgaggatcat gtgcgtggtg tacttgaaga ggcaagaaag 300
agattggggg agatcacaaa ggatcagtct cgatacagcc atgttctcca tcagttaata 360
atgcaggggc tgctgcagct cttggagccg aatgtcactg taaggacacg tgaagttgac 420
cagcgcattg ttgatagcat cttgccagct attacacaaa agtacaaaga aattactggt 480
gggaaggata tctcgctaaa agtggatact gaagccttct tgaacccaga agttactggt 540
ggaatagagt tgctagcaca aaagggcaga attaaaattg ttaatacttt ggaagcacgg 600
ctggagctca ttgcccagca acttattcct gagatacgat gtgcattgtt tggtcgtaac 660
ccaaatcgca agttcgctga ctaa 684
<210>22
<211>227
<212>PRT
<213> migratory locust (L austa migratoria)
<400>22
Met Ala Leu Ser Asp Ala Asp Val Gln Lys Gln Ile Lys His Met Met
1 5 10 15
Ala Phe Ile Asp Gln Glu Ala Asn Glu Lys Ala Glu Glu Ile Asp Ala
20 25 30
Lys Ala Glu Glu Glu Phe Asn Ile Glu Lys Gly Arg Leu Val Gln Gln
35 40 45
Gln Arg Leu Lys Ile Met Glu Tyr Tyr Glu Arg Lys Glu Lys Gln Val
50 55 60
Glu Leu Gln Lys Lys Ile Gln Ser Ser Asn Met Leu Asn Gln Ala Arg
65 70 75 80
Leu Lys Ala Leu Lys Val Arg Glu Asp His Val Arg Gly Val Leu Glu
85 90 95
Glu Ala Arg Lys Arg Leu Gly Glu Ile Thr Lys Asp Gln Ser Arg Tyr
100 105 110
Ser His Val Leu His Gln Leu Ile Met Gln Gly Leu Leu Gln Leu Leu
115 120 125
Glu ProAsn Val Thr Val Arg Thr Arg Glu Val Asp Gln Arg Ile Val
130 135 140
Asp Ser Ile Leu Pro Ala Ile Thr Gln Lys Tyr Lys Glu Ile Thr Gly
145 150 155 160
Gly Lys Asp Ile Ser Leu Lys Val Asp Thr Glu Ala Phe Leu Asn Pro
165 170 175
Glu Val Thr Gly Gly Ile Glu Leu Leu Ala Gln Lys Gly Arg Ile Lys
180 185 190
Ile Val Asn Thr Leu Glu Ala Arg Leu Glu Leu Ile Ala Gln Gln Leu
195 200 205
Ile Pro Glu Ile Arg Cys Ala Leu Phe Gly Arg Asn Pro Asn Arg Lys
210 215 220
Phe Ala Asp
225
<210>23
<211>396
<212>DNA
<213> migratory locust (L austa migratoria)
<400>23
ctaagcgatg ctgatgtcca aaaacagatt aagcatatga tggcctttat agatcaagag 60
gcaaatgaaa aggccgaaga aatagatgca aaagctgaag aagaattcaa tattgagaaa 120
ggccgattgg tgcaacagca acgtttaaaa ataatggaat attatgaaag aaaggaaaaa 180
caagtggaac tccagaagaa aattcagtct tcaaacatgc taaaccaagc acgcttaaaa 240
gctctgaaag ttcgtgagga tcatgtgcgt ggtgtacttg aagaggcaag aaagagattg 300
ggggagatca caaaggatca gtctcgatac agccatgttc tccatcagtt aataatgcag 360
gggctgctgc agctcttgga gccgaatgtc actgta 396
<210>24
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>24
taatacgact cactataggg ctaagcgatg ctgatgtcca 40
<210>25
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>25
taatacgact cactataggg tacagtgaca ttcggctcca 40
<210>26
<211>372
<212>DNA
<213> migratory locust (L austa migratoria)
<400>26
atggcattac attcggctta caaagggaaa ctcgtctctg tcatcggaga tgaggacacc 60
tgcgttggct ttcttcttgg aggagttgga gaagtcaata aaagcagaca tcccaatttc 120
atggtggtgg acaaaaacac cagtgttggt gagatagaag agtgcttcaa aagatttgtg 180
aagagagatg atattgacat catactgatt aaccagaata ttgctgagat gattcgacac 240
gtaatcgaca gccactcgca gcccgttccg tctgtcctcg agataccgtc aaaggaccat 300
ccttatgatg ccaccaaaga ttcgatatta cgtagggcaa agggaatgtt caacccagat 360
gacttcaagt aa 372
<210>27
<211>123
<212>PRT
<213> migratory locust (L austa migratoria)
<400>27
Met Ala Leu His Ser Ala Tyr Lys Gly Lys Leu Val Ser Val Ile Gly
1 5 10 15
Asp Glu Asp Thr Cys Val Gly Phe Leu Leu Gly Gly Val Gly Glu Val
20 25 30
Asn Lys Ser Arg His Pro Asn Phe Met Val Val Asp Lys Asn Thr Ser
35 40 45
Val Gly Glu Ile Glu Glu Cys Phe Lys Arg Phe Val Lys Arg Asp Asp
50 55 60
Ile Asp Ile Ile Leu Ile Asn Gln Asn Ile Ala Glu Met Ile Arg His
65 70 75 80
Val Ile Asp Ser His Ser Gln Pro Val Pro Ser Val Leu Glu Ile Pro
85 90 95
Ser Lys Asp His Pro Tyr Asp Ala Thr Lys Asp Ser Ile Leu Arg Arg
100 105 110
Ala Lys Gly Met Phe Asn Pro Asp Asp Phe Lys
115 120
<210>28
<211>297
<212>DNA
<213> migratory locust (L austa migratoria)
<400>28
acattcggct tacaaaggga aactcgtctc tgtcatcgga gatgaggaca cctgcgttgg 60
ctttcttctt ggaggagttg gagaagtcaa taaaagcaga catcccaatt tcatggtggt 120
ggacaaaaac accagtgttg gtgagataga agagtgcttc aaaagatttg tgaagagaga 180
tgatattgac atcatactga ttaaccagaa tattgctgag atgattcgac acgtaatcga 240
cagccactcg cagcccgttc cgtctgtcct cgagataccg tcaaaggacc atcctta 297
<210>29
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>29
taatacgact cactataggg acattcggct tacaaaggga 40
<210>30
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>30
taatacgact cactataggg taaggatggt cctttgacgg 40
<210>31
<211>357
<212>DNA
<213> migratory locust (L austa migratoria)
<400>31
atggctagcc agactcaggg aattcaacag cttctagcag ctgaaaaaag ggcagcagag 60
aaggtttcag aagcaagaaa gcgaaaagca cgcaggttga aacaggccaa agaagaagcc 120
caggaagaaa ttgaaaaata cagacaagaa agagagaggc agtttaaaga gtttgaagca 180
aagcacatgg gctcacgaga ggatagggct gcacagattg aagcagaaac aaaagtaaag 240
attgcagaaa tggagaaatc tgtaaatcag cggaaggaaa agttgataga aaggattttg 300
gaattggtgt acgatatcaa gcccgaactc cataagaact tcagagctgc aaagtag 357
<210>32
<211>118
<212>PRT
<213> migratory locust (L austa migratoria)
<400>32
Met Ala Ser Gln Thr Gln Gly Ile Gln Gln Leu Leu Ala Ala Glu Lys
1 5 10 15
Arg Ala Ala Glu Lys Val Ser Glu Ala Arg Lys Arg Lys Ala Arg Arg
20 25 30
Leu Lys Gln Ala Lys Glu Glu Ala Gln Glu Glu Ile Glu Lys Tyr Arg
35 40 45
Gln Glu Arg Glu Arg Gln Phe Lys Glu Phe Glu Ala Lys His Met Gly
50 55 60
Ser Arg Glu Asp ArgAla Ala Gln Ile Glu Ala Glu Thr Lys Val Lys
65 70 75 80
Ile Ala Glu Met Glu Lys Ser Val Asn Gln Arg Lys Glu Lys Leu Ile
85 90 95
Glu Arg Ile Leu Glu Leu Val Tyr Asp Ile Lys Pro Glu Leu His Lys
100 105 110
Asn Phe Arg Ala Ala Lys
115
<210>33
<211>240
<212>DNA
<213> migratory locust (L austa migratoria)
<400>33
aagggcagca gagaaggttt cagaagcaag aaagcgaaaa gcacgcaggt tgaaacaggc 60
caaagaagaa gcccaggaag aaattgaaaa atacagacaa gaaagagaga ggcagtttaa 120
agagtttgaa gcaaagcaca tgggctcacg agaggatagg gctgcacaga ttgaagcaga 180
aacaaaagta aagattgcag aaatggagaa atctgtaaat cagcggaagg aaaagttgat 240
<210>34
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>34
taatacgact cactataggg aagggcagca gagaaggttt 40
<210>35
<211>40
<212>DNA
<213> migratory locust (L austa migratoria)
<400>35
taatacgact cactataggg atcaactttt ccttccgctg 40

Claims (7)

1. The locusta migratoria V-ATPase-V1 structural domain gene is characterized in that: the gene comprises the following genes:
the nucleotide sequence of the migratory locust V-ATPase-A gene is SEQ ID NO: 1;
the nucleotide sequence of the migratory locust V-ATPase-B gene is SEQ ID NO: 6;
the nucleotide sequence of the migratory locust V-ATPase-C gene is SEQ ID NO: 11;
the nucleotide sequence of the migratory locust V-ATPase-D gene is SEQ ID NO: 16;
the nucleotide sequence of the migratory locust V-ATPase-E gene is SEQ ID NO: 21;
the nucleotide sequence of the migratory locust V-ATPase-F gene is SEQ ID NO: 26;
the nucleotide sequence of the migratory locust V-ATPase-G gene is SEQ ID NO: 31.
2. the locusta migratoria V-ATPase-V1 structural domain gene fragment is characterized in that: the following gene segments are included:
the nucleotide sequence of the migratory locust V-ATPase-A gene fragment is SEQ ID NO: 3;
the nucleotide sequence of the migratory locust V-ATPase-B gene fragment is SEQ ID NO: 8;
the nucleotide sequence of the migratory locust V-ATPase-C gene fragment is SEQ ID NO: 13;
the nucleotide sequence of the migratory locust V-ATPase-D gene fragment is SEQ ID NO: 18;
the nucleotide sequence of the migratory locust V-ATPase-E gene fragment is SEQ ID NO: 23;
the nucleotide sequence of the migratory locust V-ATPase-F gene fragment is SEQ ID NO: 28;
the nucleotide sequence of the migratory locust V-ATPase-G gene fragment is SEQ ID NO: 33.
3. the dsRNA of the locusta migratoria V-ATPase-V1 structural domain gene is characterized in that: including the following dsrnas:
the locust migratory V-ATPase-A gene fragment SEQ ID NO: 3 a synthetic dsRNA;
the locust migratory V-ATPase-B gene fragment SEQ ID NO: 8 synthetic dsRNA;
the locust-migratory V-ATPase-C gene fragment SEQ ID NO: 13 synthetic dsRNA;
the locust migratory V-ATPase-D gene fragment SEQ ID NO: 18 synthetic dsRNA;
the locust migratory V-ATPase-E gene fragment SEQ ID NO: 23 synthetic dsRNA;
the locust migratory V-ATPase-F gene fragment SEQ ID NO: 28 synthetic dsRNA;
the locust migratory V-ATPase-G gene fragment SEQ ID NO: 33 synthetic dsRNA.
4. The synthetic method of dsRNA of locusta migratoria V-ATPase-V1 structural domain gene is characterized in that: respectively carrying out gene sequence comparison on the sequences of the V-ATPase-A gene SEQ ID NO: 1. V-ATPase-B gene SEQ ID NO: 6. V-ATPase-C gene SEQ ID NO: 11. V-ATPase-D gene SEQ ID NO: 16. V-ATPase-E gene SEQ ID NO: 21. V-ATPase-F gene SEQ ID NO: 26. V-ATPase-G gene SEQ ID NO: 31 as a template, and an upstream primer and a downstream primer corresponding to the gene, and carrying out PCR amplification to obtain a DNA template with two ends provided with a T7 promoter according to HiScribeTMThe T7 High Yield RNA Synthesis Kit indicates that the corresponding dsRNA is synthesized by in vitro transcription.
5. The method for synthesizing dsRNA of locusta migratoria V-ATPase-V1 domain gene according to claim 4, wherein the method comprises the following steps:
the upstream primer and the downstream primer of the migratory locust V-ATPase-A gene are respectively as follows: SEQ ID NO: 4 and SEQ ID NO: 5;
the upstream primer and the downstream primer of the migratory locust V-ATPase-B gene are respectively as follows: SEQ ID NO: 9 and SEQ ID NO: 10;
the upstream primer and the downstream primer of the migratory locust V-ATPase-C gene are respectively as follows: SEQ ID NO: 14 and SEQ ID NO: 15;
the upstream primer and the downstream primer of the migratory locust V-ATPase-D gene are respectively as follows: SEQ ID NO: 19 and SEQ ID NO: 20;
the upstream primer and the downstream primer of the migratory locust V-ATPase-E gene are respectively as follows: SEQ ID NO: 24 and SEQ ID NO: 25;
the upstream primer and the downstream primer of the migratory locust V-ATPase-F gene are respectively as follows: SEQ ID NO: 29 and SEQ ID NO: 30, of a nitrogen-containing gas;
the upstream primer and the downstream primer of the migratory locust V-ATPase-G gene are respectively as follows: SEQ ID NO: 34 and SEQ ID NO: 35.
6. the application of dsRNA of locusta migratoria V-ATPase-V1 structural domain gene in pest control is characterized in that: the dsRNA of the V-ATPase-V1 structural domain gene of the migratory locust is applied to the control of the migratory locust, and the dsRNA is prepared into a spraying pesticide or a bait agent or is transferred into a migratory locust feeding plant body.
7. The use of dsRNA of locusta migratoria V-ATPase-V1 domain gene in pest control according to claim 6, which is characterized by: the dsRNA of the V-ATPase-V1 structural domain gene of the migratory locust comprises dsRNA of a V-ATPase-A gene, a V-ATPase-B gene, a V-ATPase-C gene, a V-ATPase-D gene, a V-ATPase-E gene, a V-ATPase-F gene and a V-ATPase-G gene, and one or more dsRNA is/are optionally prepared into a spraying pesticide or bait, or the dsRNA is transferred into a migratory locust feeding plant body to be applied to the control of the migratory locust.
CN202010249056.6A 2020-03-31 2020-03-31 Migratory locust V-ATPase-V1 structural domain gene and application of dsRNA thereof in pest control Active CN111394371B (en)

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