Method for avoiding ultraviolet degradation of dsrna bacterial solution, biological preparation for controlling tobacco whitefly and control method

Abstract

The application discloses a method for avoiding dsRNA bacterial liquid from being degraded by ultraviolet, a biological preparation for preventing and treating Bemisia tabaci and a prevention and treatment method, and belongs to the technical field of pest control. The method for avoiding the dsRNA bacterial liquid from being degraded by ultraviolet comprises the following steps: mixing the dsRNA bacterial liquid with a UV protective agent to prepare a mixed liquid of the UV protective agent and the dsRNA bacterial liquid; and the UV protective agent is one of potassium humate and sodium humate. The UV protective agent in the application can effectively avoid the degradation of the dsRNA bacterial liquid, so that the dsGawky can fully exert a fatal effect on the Bemisia tabaci and accurately achieve the goal of pest prevention and control. The method has the advantages of high efficiency, wide application, easy formula, high insecticidal efficiency, environmental friendliness and plant growth promotion, and has a good application prospect.

Description

Technical Field

[0001] This invention belongs to the field of pest control technology, specifically relating to a method for preventing dsRNA bacterial solution from being degraded by ultraviolet light, a biological agent for controlling whiteflies, and a control method. Background Technology

[0002] RNA interference (RNAi) is a mechanism triggered by short RNA fragments (siRNAs) to promote the degradation of homologous mRNAs or inhibit their translation. RNAi technology can precisely target and silence key genes in pests by specifically inhibiting gene expression, thereby achieving highly efficient pest control.

[0003] The Gawky gene plays a role in microRNA (miRNA) and short interfering RNA (siRNA)-mediated gene silencing. Restricting the expression of this gene can cause other GW body proteins to displace and impair RNAi and microRNA-induced gene silencing.

[0004] RNA interference fragments are sprayed directly onto plant surfaces, allowing pests to ingest or absorb these fragments. However, directly sprayed double-stranded dsRNA bacterial solutions are susceptible to degradation by nucleases and ultraviolet light in the air, preventing effective triggering of the RNAi reaction and thus weakening its interference effect on pests. Therefore, exploring strategies to avoid UV degradation of dsRNA bacterial solutions is crucial for promoting the application of RNAi technology in the field of biological pest control. Summary of the Invention

[0005] In view of the problems existing in the prior art, the purpose of this invention is to provide a method to prevent dsRNA bacterial solution from being degraded by ultraviolet light, a biological agent for controlling whiteflies, and a control method.

[0006] To achieve the above objectives, the present invention adopts the following technical solution:

[0007] A method for preventing dsRNA bacterial culture from being degraded by ultraviolet light involves mixing the dsRNA bacterial culture with a UV protectant to prepare a mixture of UV protectant and dsRNA bacterial culture; wherein the UV protectant is one of potassium humate or sodium humate.

[0008] The above-mentioned method for preventing the degradation of dsRNA bacterial culture by ultraviolet light is applied to the control of agricultural pests using RNA interference.

[0009] A biological agent for controlling whiteflies comprises a UV protectant and a dsRNA bacterial suspension that interferes with the Gawky gene of whiteflies; the UV protectant is one of potassium humate and sodium humate.

[0010] Based on the above scheme, the bacterial solution that interferes with the whitefly Gawky gene dsRNA is a bacterial cell solution that transcribes the whitefly Gawky gene dsRNA.

[0011] Based on the above scheme, the bacterial cell solution for transcribing the Gawky gene dsRNA of the whitefly is prepared by the following method:

[0012] The dsRNA sequence of the Gawky gene was amplified using primers and ligated into an expression vector to construct a recombinant vector, which was then transformed into the expression strain. The expression strain was induced to transcribe the dsRNA sequence of the Gawky gene. The supernatant of the bacterial culture was discarded, and the culture was dissolved and diluted with ddH2O to obtain the final product.

[0013] Based on the above scheme, the method for inducing the expression strain to transcribe the Gawky gene dsRNA sequence is as follows:

[0014] Recombinant bacteria transformed with a recombinant vector expressing the Gawky gene dsRNA from the whitefly were inoculated into LB medium containing Amp and cultured at 37°C with shaking at 20 rpm. When the bacterial culture OD... 600 When the value reaches 0.5, add IPTG solution with a final concentration of 0.5M and induce for 4 hours to obtain the final product.

[0015] Based on the above scheme, the preparation method of the biological agent for controlling whiteflies is as follows:

[0016] (1) Disperse the UV protectant in 0.1% Tween80 aqueous solution to prepare a UV protectant solution with a concentration of 0.2%-5%;

[0017] (2) The UV protectant solution and the bacterial cell solution that transcribes the Gawky gene dsRNA of the whitefly are mixed at a volume ratio of 1:1 and shaken to mix well to obtain a biological agent for the control of whiteflies.

[0018] The application of the above-mentioned biological agents for controlling whiteflies in the biological control of whiteflies.

[0019] A biological control method for tobacco whiteflies involves spraying plants with the aforementioned biological agent for controlling tobacco whiteflies, causing the whiteflies to feed on the sprayed plants, thereby increasing the mortality rate of the tobacco whiteflies.

[0020] Based on the above scheme, the whitefly mentioned is either type Q or type B.

[0021] Advantages of the technical solution of this invention

[0022] This invention develops a technology based on the UV protectant KH / SH to prevent ultraviolet radiation from degrading dsRNA bacterial suspensions. The process involves shaking and mixing a humate solution with the dsRNA bacterial suspension to form a mixture of UV protectant (KH or SH) and dsRNA. The UV protectant safeguards the dsRNA bacterial suspension from UV degradation, thereby achieving the lethal effect of dsGawky on whiteflies and achieving pest control. This material effectively protects the dsRNA bacterial suspension by absorbing ultraviolet radiation through its specific organic molecules, reducing direct damage from ultraviolet radiation. This method is highly efficient, widely applicable, easy to formulate, has high insecticidal efficacy, is environmentally friendly, and promotes plant growth, showing great promise for future applications. Attached Figure Description

[0023] Figure 1 The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0024] Figure 2 The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0025] Figure 3 Line graph showing the lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0026] Figure 4 The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed from dsGawky bacteria on Q-type whitefly eggs;

[0027] Figure 5 The lethal effect of outdoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed dsGawky bacteria on Q-type whitefly nymphs.

[0028] Figure 6 The lethality of adult Q-type whiteflies was determined by spraying a bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed dsGawky bacteria under indoor ultraviolet light irradiation for 2 hours.

[0029] Figure 7 Line graph showing the lethal effect of spraying a bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed dsGawky bacteria on adult Q-type whiteflies under indoor ultraviolet light irradiation for 2 hours.

[0030] Figure 8 The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0031] Figure 9 The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0032] Figure 10 Line graph showing the lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0033] Figure 11 The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed from dsGawky bacteria on Q-type whitefly eggs;

[0034] Figure 12 The lethal effect of outdoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed dsGawky bacteria on Q-type whitefly nymphs.

[0035] Figure 13 The lethality of adult Q-type whiteflies was determined by spraying a bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed dsGawky bacteria under indoor ultraviolet light irradiation for 2 hours.

[0036] Figure 14 Line graph showing the lethal effect of spraying a bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed dsGawky bacteria on adult Q-type whiteflies under indoor ultraviolet light irradiation for 2 hours.

[0037] Figure 15 The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 0.2% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies;

[0038] Figure 16 The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 0.2% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies;

[0039] Figure 17 The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 5% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies;

[0040] Figure 18 The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 5% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0041] Figure 19 The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 0.2% UV protectant SH and transcribed from dsGawky bacteria on adult Q-type whiteflies;

[0042] Figure 20 The lethality of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 0.2% UV protectant SH and transcribed from dsGawky bacteria against adult Q-type whiteflies;

[0043] Figure 21 The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 5% UV protectant SH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0044] Figure 22 The lethality of adult Q-type whiteflies by outdoor spraying of bacterial solutions transcribed from dsGawky bacteria or a mixture of 5% UV protectant SH and transcribed dsGawky bacteria. Detailed Implementation

[0045] The terminology used in this invention, unless otherwise specified, generally has the meanings commonly understood by those skilled in the art. The invention is further described in detail below with reference to specific embodiments and data. The following embodiments are merely illustrative and are not intended to limit the scope of the invention in any way.

[0046] Unless otherwise specified, the experimental methods used in the following embodiments are conventional methods, performed according to the techniques or conditions described in the literature in this field or according to the product instructions. Unless otherwise specified, the experimental materials, reagents, and chemicals used in the following embodiments can be obtained through general channels.

[0047] The whitefly populations used in the following examples were collected in January 2017 from Lingshui City, Hainan Province. These insects were kept on common tobacco variety NC89 in insect-proof cages for an extended period (temperature 27±1℃, relative humidity 60%±5%, photoperiod 16L:8D).

[0048] Example 1

[0049] Application of potassium humate in preventing dsRNA bacterial culture from being degraded by ultraviolet light

[0050] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, potassium humate was prepared into a 1% (mass fraction) solution as a 1% UV protectant KH solution.

[0051] (2) Mixing the UV protectant KH solution with the dsRNA bacterial solution at a volume ratio of 1:1 can prevent the dsRNA bacterial solution from being degraded by ultraviolet light.

[0052] Example 2

[0053] A method to prevent the degradation of whitefly dsRNA bacterial culture by ultraviolet light.

[0054] (1) Disperse 0.2 mL of Tween80 in 199.8 mL of water and stir until homogeneous to obtain a 0.1% Tween80 solution as a solvent;

[0055] (2) Take 1g of potassium humate and disperse it in 99mL of the 0.1% Tween80 solution prepared in step (1). Stir thoroughly for 30min until the solution becomes uniform, and you will get a 1% UV protectant KH solution.

[0056] (3) Mix the 1% UV protectant KH solution obtained in step (2) with the whitefly dsRNA bacterial solution at a volume ratio of 1:1 to avoid the whitefly dsRNA bacterial solution being degraded by ultraviolet light.

[0057] Example 3

[0058] The biological control method for whiteflies based on 1% UV protectant KH consists of the following steps:

[0059] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, potassium humate was prepared into a 1% (mass fraction) solution as UV protectant KH solution.

[0060] (2) Mix 1% UV protectant KH solution with a solution of dsGawky cells (i.e., cells that transcribe the whitefly Gawky gene dsRNA) (concentration 2 g / L) at a volume ratio of 1:1. After shaking and mixing, a mixture of 1% UV protectant KH and dsGawky cells for whitefly RNAi interference is obtained.

[0061] (3) The above-mentioned 1% UV protectant KH and the transcribed dsGawky bacteria were sprayed on the plants, causing the whiteflies to feed on the sprayed plants, thereby increasing the mortality rate of the whiteflies.

[0062] The method for preparing the solution transcribed from dsGawky cells is as follows:

[0063] Based on the gene sequence of the whitefly Gawky (SEQ ID NO.1), primers (SEQ ID NO.2 and SEQ ID NO.3) for Gawky gene dsRNA were designed and prepared. PCR amplification was performed using cDNA reverse transcribed from total whitefly RNA as a template. The PCR reaction system consisted of 2.5 μL of 10x buffer, 2 μL of dNTPs, 1 μL of upstream primer, 1 μL of downstream primer, 0.25 μL of rTaq, 17.25 μL of H2O, and 1 μL of cDNA. The reaction procedure was as follows: pre-denaturation at 90℃ for 15 min; denaturation at 95℃ for 10 s, annealing at 60℃ for 30 s, 72℃ for 10 s, for 40 cycles; extension PCR at 72℃ for 10 min. After the PCR was completed, the product was recovered, and the amplified product was ligated into the linearized L4440 plasmid using T4 DNA ligase. The plasmid was then transformed into HT115(DE3) competent cells, and finally 300 μL was plated onto a plate containing 100 mg / mL Ampicillin. + The culture was incubated overnight at 37°C on a solid medium, then inverted. The next day, six single colonies were randomly selected from the plates, dissolved in the medium, and cultured to prepare a bacterial suspension. The bacteria that were successfully identified by PCR were the target recombinant bacteria.

[0064] The recombinant bacteria were inoculated into 15 mL of LB medium (Amp). + The culture was incubated at 37℃ with shaking at 20 rpm. When the OD value of the bacterial culture reached 0.5, 0.5 M IPTG solution was added for induction for 4 h to obtain the transcriptional dsGawky bacterial culture. The bacterial culture was centrifuged at 4℃ and 5000×g for 10 min to separate the bacterial cells. The supernatant was discarded, and the bacterial cells were resuspended in a small amount of PBS solution, mixed well, and centrifuged at 4℃ and 5000×g for 5 min. The supernatant was discarded, and the process was repeated twice. The bacterial cells were stored at -20℃ for later use.

[0065] When using, dissolve and dilute with ddH2O to obtain a bacterial solution (wet bacterial cells) concentration of 2g / L.

[0066] SEQ ID NO.1 (5'→3'):

[0067]

[0068] The primers used to prepare Gawky gene dsRNA are as follows:

[0069] Forward primer: dsGawkyF:5'-taatacgactcactatagggGGAGATCCGTCCTACACGCTACA-3' (SEQ ID NO.2);

[0070] Reverse primer: dsGawkyR:5'-taatacgactcactatagggCGAGTTGGTGGAGGTTGCTGAAG-3' (SEQ ID NO.3).

[0071] Example 4

[0072] The lethal effect of indoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed dsGawky bacteria on adult Q-type whiteflies.

[0073] The transcribed dsGawky bacterial cell solution prepared in Example 3 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator (no sunlight) at 27 ± 1 °C, relative humidity 60% ± 5%, and a photoperiod of 16 L:8 D. Adult mortality was recorded 7 days after spraying.

[0074] The mixture of 1% UV protectant KH prepared in Example 3 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator with no sunlight, a temperature of 27±1℃, a relative humidity of 60%±5%, and a photoperiod of 16L:8D. Adult mortality was recorded 7 days after spraying.

[0075] The results are as follows Figure 1 As shown, under indoor conditions, the mortality rate of adult Q-type whiteflies was 53.9% when sprayed with a solution of transcribed dsGawky bacteria, and the mortality rate of adult Q-type whiteflies was 68.3% when sprayed with a mixture of 1% UV protectant KH and transcribed dsGawky bacteria.

[0076] Example 5

[0077] The lethal effect of outdoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed dsGawky bacteria on adult Q-type whiteflies.

[0078] The transcribed dsGawky bacterial cell solution prepared in Example 3 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed outdoors in sunlight. Adult mortality rates were recorded 7 days after spraying.

[0079] The mixture of 1% UV protectant KH prepared in Example 3 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates for each treatment. The plants were placed outdoors in sunlight. The adult mortality rate was recorded 7 days after spraying.

[0080] The results are as follows Figure 2 and Figure 3 As shown, under outdoor conditions, the mortality rate of adult Q-type whiteflies was 18% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of adult Q-type whiteflies was 42.8% when sprayed with a mixture of 1% UV protectant KH and transcribed dsGawky bacteria. The two solutions showed a significant difference.

[0081] Example 6

[0082] The lethal effect of outdoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed dsGawky bacteria on Q-type whitefly eggs.

[0083] Fifteen pairs of whiteflies were released onto cotton plants. One day after the whiteflies laid eggs, the adults were removed, and the number of eggs on each cotton plant was recorded under a microscope. The dsGawky bacterial cell solution prepared in Example 3 was dissolved and diluted using ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto the cotton plants, three times on each side of the leaves, with three replicates for each treatment. The plants were placed outdoors in sunlight. Eggs that did not hatch into nymphs by day 10 after spraying were considered dead, and the egg mortality rate was recorded.

[0084] Fifteen pairs of whiteflies were released onto cotton plants. One day after the whiteflies laid eggs, the adults were removed, and the number of eggs on each cotton plant was recorded under a microscope. A mixture of 1% UV protectant KH prepared in Example 3 and transcribed dsGawky bacteria was sprayed onto the cotton plants, three times on each side of the leaves, with three replicates for each treatment. The plants were placed outdoors in sunlight. Eggs that did not hatch into nymphs by day 10 after spraying were considered dead, and the egg mortality rate was recorded.

[0085] The results are as follows Figure 4As shown, under outdoor conditions, the lethality of spraying a solution of transcribed dsGawky bacteria against Q-type whitefly eggs was 18%, while the lethality of spraying a mixture of 1% UV protectant KH and transcribed dsGawky bacteria against Q-type whitefly eggs was 41.7%, showing a significant difference between the two.

[0086] Example 7

[0087] The lethal effect of outdoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed dsGawky bacteria on Q-type whitefly nymphs.

[0088] Fifteen pairs of whiteflies were released onto cotton plants. On day 9, the number of nymphs was observed under a microscope. The transcribed dsGawky bacterial solution prepared in Example 3 was dissolved and diluted using ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto the cotton plants, three times on each side of the leaves. Each treatment was repeated three times, and the plants were placed outdoors in sunlight. Because the emergence time of whitefly nymphs varied slightly, some nymphs had already emerged as adults by day 17. Since it was not possible to accurately determine nymph mortality under a microscope, newly emerged adults were collected and their numbers recorded daily from day 17. By day 25, no newly emerged adults had appeared for three consecutive days. At this point, the unemerged nymphs were considered dead, and the nymph mortality rate was calculated.

[0089] Fifteen pairs of whiteflies were released onto cotton plants. On day 9, the number of nymphs was observed under a microscope. A mixture of 1% UV protectant KH prepared in Example 3 and transcribed dsGawky bacteria was sprayed onto the cotton plants, three times on each side of the leaves, with three replicates for each treatment. The plants were placed outdoors in sunlight. Because the emergence time of whitefly nymphs varied slightly, some nymphs had already emerged as adults by day 17. Since microscopic examination could not accurately determine nymph mortality, newly emerged adults were collected and their numbers recorded daily from day 17. By day 25, no newly emerged adults had appeared for three consecutive days. At this point, the unemerged nymphs were considered dead, and the nymph mortality rate was calculated.

[0090] The results are as follows Figure 5 As shown, under outdoor conditions, the mortality rate of Q-type whitefly nymphs was 22% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of Q-type whitefly nymphs was 50.4% when sprayed with a mixture of 1% UV protectant KH and transcribed dsGawky bacteria. The two methods showed a significant difference.

[0091] Example 8

[0092] Under indoor ultraviolet light irradiation for 2 hours, the lethal effect of spraying a bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant KH and transcribed dsGawky bacteria on adult Q-type whiteflies was investigated.

[0093] The transcribed dsGawky bacterial cell solution prepared in Example 3 was dissolved and diluted using ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After irradiation with ultraviolet light for 2 hours and allowing the solution to dry, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator at 27 ± 1℃, relative humidity 60% ± 5%, and a photoperiod of 16 L:8 D. Adult mortality was recorded 7 days after spraying.

[0094] The mixture of 1% UV protectant KH prepared in Example 3 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the leaves. After irradiation with UV light for 2 hours and allowing the solution to dry, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator at 27±1℃, 60%±5% relative humidity, and a photoperiod of 16L:8D. Adult mortality was recorded 7 days after spraying.

[0095] The results are as follows Figure 6 and Figure 7 As shown, under 2 hours of UV irradiation, the mortality rate of adult Q-type whiteflies was 30.1% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of adult Q-type whiteflies was 43.4% when sprayed with a mixture of UV protectant KH and transcribed dsGawky bacteria. The two results showed a significant difference.

[0096] Example 9

[0097] Application of sodium humate in preventing dsRNA bacterial culture from being degraded by ultraviolet light

[0098] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, sodium humate was prepared into a 1% (mass fraction) solution as a 1% UV protectant SH solution.

[0099] (2) Mixing 1% UV protectant SH solution with dsRNA bacterial solution at a volume ratio of 1:1 can prevent dsRNA bacterial solution from being degraded by UV.

[0100] Example 10

[0101] A method to prevent the degradation of whitefly dsRNA bacterial culture by ultraviolet light.

[0102] (1) Disperse 0.2 mL of Tween80 in 199.8 mL of water and stir until homogeneous to obtain a 0.1% Tween80 solution as a solvent;

[0103] (2) Take 1g of sodium humate and disperse it in 99mL of the 0.1% Tween80 solution prepared in step (1). Stir thoroughly for 30min until the solution becomes uniform, and you will get a 1% UV protectant SH solution.

[0104] (3) Mix the 1% UV protectant SH solution obtained in step (2) with the whitefly dsRNA bacterial solution at a volume ratio of 1:1 to avoid the whitefly dsRNA bacterial solution being degraded by ultraviolet light.

[0105] Example 11

[0106] The biological control method for whiteflies based on 1% UV protectant SH consists of the following steps:

[0107] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, sodium humate was prepared into a 1% (mass fraction) solution as a 1% UV protectant SH solution.

[0108] (2) Mix 1% UV protectant SH solution with a solution of dsGawky cells (i.e., cells that transcribe the whitefly Gawky gene dsRNA) (concentration 2 g / L) at a volume ratio of 1:1. After shaking and mixing, a mixture of 1% UV protectant SH and dsGawky cells for whitefly RNAi interference is obtained.

[0109] (3) The above-mentioned UV protectant SH and the transcribed dsGawky bacteria were sprayed on the plants, causing the whiteflies to feed on the sprayed plants, thereby increasing the mortality rate of the whiteflies.

[0110] The method for preparing the solution for transcribing dsGawky cells is the same as in Example 3.

[0111] Example 12

[0112] The lethal effect of indoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed dsGawky bacteria on adult Q-type whiteflies.

[0113] The transcribed dsGawky bacterial cell solution prepared in Example 11 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator (no sunlight) at 27 ± 1°C, relative humidity 60% ± 5%, and a photoperiod of 16 L:8 D. Adult mortality was recorded 7 days after spraying.

[0114] The mixture of 1% UV protectant SH prepared in Example 11 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator with no sunlight, a temperature of 27±1℃, a relative humidity of 60%±5%, and a photoperiod of 16L:8D. Adult mortality was recorded 7 days after spraying.

[0115] The results are as follows Figure 8 As shown, under indoor conditions, the mortality rate of adult Q-type whiteflies was 53.9% when sprayed with a solution of transcribed dsGawky bacteria, and the mortality rate of adult Q-type whiteflies was 62.7% when sprayed with a mixture of 1% UV protectant SH and transcribed dsGawky bacteria.

[0116] Example 13

[0117] The lethal effect of outdoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed dsGawky bacteria on adult Q-type whiteflies.

[0118] The transcribed dsGawky bacterial cell solution prepared in Example 11 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed outdoors in sunlight. Adult mortality rates were recorded 7 days after spraying.

[0119] The mixture of 1% UV protectant SH prepared in Example 11 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates for each treatment. The plants were placed outdoors in sunlight. The adult mortality rate was recorded 7 days after spraying.

[0120] The results are as follows Figure 9 and Figure 10 As shown, under outdoor conditions, the mortality rate of adult Q-type whiteflies was 18% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of adult Q-type whiteflies was 48% when sprayed with a mixture of 1% UV protectant SH and transcribed dsGawky bacteria. The two methods showed a significant difference.

[0121] Example 14

[0122] The lethal effect of outdoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed dsGawky bacteria on Q-type whitefly eggs.

[0123] Fifteen pairs of whiteflies were released onto cotton plants. One day after the whiteflies laid eggs, the adults were removed, and the number of eggs on each cotton plant was recorded under a microscope. The dsGawky bacterial cell solution prepared in Example 11 was dissolved and diluted using ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto the cotton plants, three times on each side of the leaves, with three replicates for each treatment. The plants were placed outdoors in sunlight. Eggs that did not hatch into nymphs by day 10 after spraying were considered dead, and the egg mortality rate was recorded.

[0124] Fifteen pairs of whiteflies were released onto cotton plants. One day after the whiteflies laid eggs, the adults were removed, and the number of eggs on each cotton plant was recorded under a microscope. A mixture of 1% UV protectant SH prepared in Example 11 and transcribed dsGawky bacteria was sprayed onto the cotton plants, three times on each side of the leaves, with three replicates for each treatment. The plants were placed outdoors in sunlight. Eggs that did not hatch into nymphs by day 10 after spraying were considered dead, and the egg mortality rate was recorded.

[0125] The results are as follows Figure 11 As shown, under outdoor conditions, the lethality of spraying a solution of transcribed dsGawky bacteria against Q-type whitefly eggs was 18%, while the lethality of spraying a mixture of 1% UV protectant SH and transcribed dsGawky bacteria against Q-type whitefly eggs was 32%, showing a significant difference between the two.

[0126] Example 15

[0127] The lethal effect of outdoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed dsGawky bacteria on Q-type whitefly nymphs.

[0128] Fifteen pairs of whiteflies were released onto cotton plants. On day 9, the number of nymphs was observed under a microscope. The transcribed dsGawky bacterial solution prepared in Example 11 was dissolved and diluted using ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto the cotton plants, three times on each side of the leaves. Each treatment was repeated three times, and the plants were placed outdoors in sunlight. Because the emergence time of whitefly nymphs varied slightly, some nymphs had already emerged as adults by day 17. Since it was not possible to accurately determine nymph mortality under a microscope, newly emerged adults were collected and their numbers recorded daily from day 17. By day 25, no newly emerged adults had appeared for three consecutive days. At this point, the unemerged nymphs were considered dead, and the nymph mortality rate was calculated.

[0129] Fifteen pairs of whiteflies were released onto cotton plants. On day 9, the number of nymphs was observed under a microscope. A mixture of the UV protectant SH prepared in Example 11 and the transcribed dsGawky bacteria was sprayed onto the cotton plants, three times on each side of the leaves, with three replicates for each treatment. The plants were placed outdoors in sunlight. Because the emergence time of whitefly nymphs varied slightly, some nymphs had already emerged as adults by day 17. Since microscopic examination could not accurately determine nymph mortality, newly emerged adults were collected and their numbers recorded daily from day 17. By day 25, no newly emerged adults had appeared for three consecutive days. At this point, the unemerged nymphs were considered dead, and the nymph mortality rate was calculated.

[0130] The results are as follows Figure 12 As shown, under outdoor conditions, the mortality rate of Q-type whitefly nymphs was 22% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of Q-type whitefly nymphs was 46.8% when sprayed with a mixture of 1% UV protectant SH and transcribed dsGawky bacteria. The two methods showed a significant difference.

[0131] Example 16

[0132] Under indoor ultraviolet light irradiation for 2 hours, the lethal effect of spraying a bacterial solution transcribed from dsGawky bacteria or a mixture of 1% UV protectant SH and transcribed dsGawky bacteria on adult Q-type whiteflies was investigated.

[0133] The transcribed dsGawky bacterial cell solution prepared in Example 11 was dissolved and diluted using ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After irradiation with ultraviolet light for 2 hours and allowing the solution to dry, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator at 27 ± 1℃, relative humidity 60% ± 5%, and a photoperiod of 16 L:8 D. Adult mortality rates were recorded 7 days after spraying.

[0134] The mixture of 1% UV protectant SH prepared in Example 11 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the leaves. After irradiation with UV light for 2 hours and allowing the solution to dry, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator at 27±1℃, 60%±5% relative humidity, and a photoperiod of 16L:8D. Adult mortality was recorded 7 days after spraying.

[0135] The results are as follows Figure 13 and Figure 14As shown, under 2 hours of UV irradiation, the mortality rate of adult Q-type whiteflies was 30.1% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of adult Q-type whiteflies was 42.8% when sprayed with a mixture of 1% UV protectant SH and transcribed dsGawky bacteria. The two results showed a significant difference.

[0136] Example 17

[0137] Application of potassium humate in preventing dsRNA bacterial culture from being degraded by ultraviolet light

[0138] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, potassium humate was prepared into a 0.2% (mass fraction) solution as a 0.2% UV protectant KH solution.

[0139] (2) Mixing 0.2% UV protectant KH solution with dsRNA bacterial solution at a volume ratio of 1:1 can prevent dsRNA bacterial solution from being degraded by UV.

[0140] Example 18

[0141] A method to prevent the degradation of whitefly dsRNA bacterial culture by ultraviolet light.

[0142] (1) Disperse 0.2 mL of Tween80 in 199.8 mL of water and stir until homogeneous to obtain a 0.1% Tween80 solution as a solvent;

[0143] (2) Take 0.2g of potassium humate and disperse it in 99.8mL of the 0.1% Tween80 solution prepared in step (1). Stir thoroughly for 30min until the solution becomes uniform, and then obtain the 0.2% UV protectant KH solution.

[0144] (3) The 0.2% UV protectant KH solution obtained in step (2) is mixed with the whitefly dsRNA bacterial solution at a volume ratio of 1:1 to avoid the whitefly dsRNA bacterial solution being degraded by ultraviolet light.

[0145] Example 19

[0146] The biological control method for whiteflies based on 0.2% UV protectant KH consists of the following steps:

[0147] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, potassium humate was prepared into a 0.2% (mass fraction) solution as a 0.2% UV protectant KH solution.

[0148] (2) Mix 0.2% UV protectant KH solution with a solution of dsGawky cells (i.e., cells that transcribe the whitefly Gawky gene dsRNA) (concentration 2 g / L) at a volume ratio of 1:1. After shaking and mixing, a mixture of 0.2% UV protectant KH and dsGawky cells for whitefly RNAi interference is obtained.

[0149] (3) The above-mentioned 0.2% UV protectant KH and the transcribed dsGawky bacteria were sprayed on the plants, causing the whiteflies to feed on the sprayed plants, thereby increasing the mortality rate of the whiteflies.

[0150] The method for preparing the solution for transcribing dsGawky cells is the same as in Example 3.

[0151] Example 20

[0152] The lethal effect of indoor spraying of bacterial suspension transcribed from dsGawky bacteria or a mixture of 0.2% UV protectant KH and transcribed dsGawky bacteria on adult Q-type whiteflies.

[0153] The transcribed dsGawky bacterial cell solution prepared in Example 19 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator (no sunlight) at 27 ± 1 °C, relative humidity 60% ± 5%, and a photoperiod of 16 L:8 D. Adult mortality was recorded 7 days after spraying.

[0154] The mixture of 0.2% UV protectant KH prepared in Example 19 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator with no sunlight, a temperature of 27±1℃, a relative humidity of 60%±5%, and a photoperiod of 16L:8D. Adult mortality was recorded 7 days after spraying.

[0155] The results are as follows Figure 15 As shown, under indoor conditions, the mortality rate of adult Q-type whiteflies was 53.9% when sprayed with a solution of transcribed dsGawky bacteria, and 51.1% when sprayed with a mixture of 0.2% UV protectant KH and transcribed dsGawky bacteria.

[0156] Example 21

[0157] The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 0.2% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies;

[0158] The transcribed dsGawky bacterial cell solution prepared in Example 19 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed outdoors in sunlight. Adult mortality rates were recorded 7 days after spraying.

[0159] The mixture of 0.2% UV protectant KH prepared in Example 19 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates for each treatment. The plants were placed outdoors in sunlight. The adult mortality rate was recorded 7 days after spraying.

[0160] The results are as follows Figure 16 As shown, under outdoor conditions, the mortality rate of adult Q-type whiteflies was 18% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of adult Q-type whiteflies was 32.4% when sprayed with a mixture of 0.2% UV protectant KH and transcribed dsGawky bacteria. The two solutions showed a significant difference.

[0161] Example 22

[0162] Application of potassium humate in preventing dsRNA bacterial culture from being degraded by ultraviolet light

[0163] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, potassium humate was prepared into a 5% (mass fraction) solution as a 5% UV protectant KH solution.

[0164] (2) Mixing 5% UV protectant KH solution with dsRNA bacterial solution at a volume ratio of 1:1 can prevent dsRNA bacterial solution from being degraded by ultraviolet light.

[0165] Example 23

[0166] A method to prevent the degradation of whitefly dsRNA bacterial culture by ultraviolet light.

[0167] (1) Disperse 0.2 mL of Tween80 in 199.8 mL of water and stir until homogeneous to obtain a 0.1% Tween80 solution as a solvent;

[0168] (2) Take 5g of potassium humate and disperse it in 95mL of the 0.1% Tween80 solution prepared in step (1). Stir thoroughly for 30min until the solution becomes uniform, and you will get a 5% UV protectant KH solution.

[0169] (3) Mix the 5% UV protectant KH solution obtained in step (2) with the whitefly dsRNA bacterial solution at a volume ratio of 1:1 to avoid the whitefly dsRNA bacterial solution being degraded by ultraviolet light.

[0170] Example 24

[0171] The biological control method for whiteflies based on 5% UV protectant KH consists of the following steps:

[0172] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, potassium humate was prepared into a 5% (mass fraction) solution as a 5% UV protectant KH solution.

[0173] (2) Mix the 5% UV protectant KH solution with the bacterial cells that transcribe dsGawky (i.e., bacterial cells that transcribe the whitefly Gawky gene dsRNA) solution (concentration of 2 g / L) at a volume ratio of 1:1. After shaking and mixing, the mixture is used for whitefly RNAi interference.

[0174] (3) The above-mentioned 5% UV protectant KH and the transcribed dsGawky bacteria were sprayed on the plants, causing the whiteflies to feed on the sprayed plants, thereby increasing the mortality rate of the whiteflies.

[0175] The method for preparing the solution for transcribing dsGawky cells is the same as in Example 3.

[0176] Example 25

[0177] The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 5% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies;

[0178] The transcribed dsGawky bacterial cell solution prepared in Example 24 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator (no sunlight) at 27 ± 1 °C, relative humidity 60% ± 5%, and a photoperiod of 16 L:8 D. Adult mortality was recorded 7 days after spraying.

[0179] The mixture of 5% UV protectant KH prepared in Example 24 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator with no sunlight, a temperature of 27±1℃, a relative humidity of 60%±5%, and a photoperiod of 16L:8D. Adult mortality was recorded 7 days after spraying.

[0180] The results are as follows Figure 17 As shown, under indoor conditions, the mortality rate of adult Q-type whiteflies was 53.9% when sprayed with a solution of transcribed dsGawky bacteria, and the mortality rate of adult Q-type whiteflies was 54.9% when sprayed with a mixture of 5% UV protectant KH and transcribed dsGawky bacteria.

[0181] Example 26

[0182] The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 5% UV protectant KH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0183] The transcribed dsGawky bacterial cell solution prepared in Example 24 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed outdoors in sunlight. Adult mortality rates were recorded 7 days after spraying.

[0184] The mixture of 5% UV protectant KH prepared in Example 24 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates for each treatment. The plants were placed outdoors in sunlight. The adult mortality rate was recorded 7 days after spraying.

[0185] The results are as follows Figure 18 As shown, under outdoor conditions, the mortality rate of adult Q-type whiteflies was 18% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of adult Q-type whiteflies was 59.6% when sprayed with a mixture of 5% UV protectant KH and transcribed dsGawky bacteria. The two solutions showed a significant difference.

[0186] Example 27

[0187] Application of sodium humate in preventing dsRNA bacterial culture from being degraded by ultraviolet light

[0188] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, sodium humate was prepared into a 0.2% (mass fraction) solution as 0.2% UV protectant SH solution.

[0189] (2) Mixing 0.2% UV protectant SH solution with dsRNA bacterial solution at a volume ratio of 1:1 can prevent dsRNA bacterial solution from being degraded by UV.

[0190] Example 28

[0191] A method to prevent the degradation of whitefly dsRNA bacterial culture by ultraviolet light.

[0192] (1) Disperse 0.2 mL of Tween80 in 199.8 mL of water and stir until homogeneous to obtain a 0.1% Tween80 solution as a solvent;

[0193] (2) Take 0.2g of sodium humate and disperse it in 99.8mL of the 0.1% Tween80 solution prepared in step (1). Stir thoroughly for 30min until the solution becomes uniform, and then obtain the 0.2% UV protectant SH solution.

[0194] (3) Mix the 0.2% UV protectant SH solution obtained in step (2) with the whitefly dsRNA bacterial solution at a volume ratio of 1:1 to avoid the whitefly dsRNA bacterial solution being degraded by ultraviolet light.

[0195] Example 29

[0196] The biological control method for whiteflies based on 0.2% UV protectant SH consists of the following steps:

[0197] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, sodium humate was prepared into a 0.2% (mass fraction) solution as a 1% UV protectant SH solution.

[0198] (2) Mix 0.2% UV protectant SH solution with a solution of dsGawky cells (i.e., cells that transcribe the whitefly Gawky gene dsRNA) (concentration 2 g / L) at a volume ratio of 1:1. After shaking and mixing, the mixture of 0.2% UV protectant SH and dsGawky cells for whitefly RNAi interference is obtained.

[0199] (3) The above-mentioned 0.2% UV protectant SH and the transcribed dsGawky bacteria were sprayed on the plants, causing the whiteflies to feed on the sprayed plants, thereby increasing the mortality rate of the whiteflies.

[0200] The method for preparing the solution for transcribing dsGawky cells is the same as in Example 3.

[0201] Example 30

[0202] The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 0.2% UV protectant SH and transcribed from dsGawky bacteria on adult Q-type whiteflies;

[0203] The transcribed dsGawky bacterial cell solution prepared in Example 29 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator (no sunlight) at 27 ± 1 °C, relative humidity 60% ± 5%, and a photoperiod of 16 L:8 D. Adult mortality was recorded 7 days after spraying.

[0204] The mixture of 0.2% UV protectant SH prepared in Example 29 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator with no sunlight, a temperature of 27±1℃, a relative humidity of 60%±5%, and a photoperiod of 16L:8D. Adult mortality was recorded 7 days after spraying.

[0205] The results are as follows Figure 19 As shown, under indoor conditions, the mortality rate of adult Q-type whiteflies was 53.9% when sprayed with a solution of transcribed dsGawky bacteria, and 49.5% when sprayed with a mixture of 0.2% UV protectant SH and transcribed dsGawky bacteria.

[0206] Example 31

[0207] The lethality of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 0.2% UV protectant SH and transcribed from dsGawky bacteria against adult Q-type whiteflies;

[0208] The transcribed dsGawky bacterial cell solution prepared in Example 29 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed outdoors in sunlight. Adult mortality rates were recorded 7 days after spraying.

[0209] The mixture of 0.2% UV protectant SH prepared in Example 29 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates for each treatment. The plants were placed outdoors in sunlight. The adult mortality rate was recorded 7 days after spraying.

[0210] The results are as follows Figure 20 As shown, under outdoor conditions, the mortality rate of adult Q-type whiteflies was 18% when sprayed with a solution of transcribed dsGawky bacteria, and 33% when sprayed with a mixture of 0.2% UV protectant SH and transcribed dsGawky bacteria. The two results showed a significant difference.

[0211] Example 32

[0212] Application of sodium humate in preventing dsRNA bacterial culture from being degraded by ultraviolet light

[0213] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, sodium humate was prepared into a 5% (mass fraction) solution as a 5% UV protectant SH solution.

[0214] (2) Mixing 5% UV protectant SH solution with dsRNA bacterial solution at a volume ratio of 1:1 can prevent dsRNA bacterial solution from being degraded by UV.

[0215] Example 33

[0216] A method to prevent the degradation of whitefly dsRNA bacterial culture by ultraviolet light.

[0217] (1) Disperse 0.2 mL of Tween80 in 199.8 mL of water and stir until homogeneous to obtain a 0.1% Tween80 solution as a solvent;

[0218] (2) Take 5g of sodium humate and disperse it in 95mL of the 0.1% Tween80 solution prepared in step (1). Stir thoroughly for 30min until the solution becomes uniform, and you will get a 5% UV protectant SH solution.

[0219] (3) Mix the 5% UV protectant SH solution obtained in step (2) with the whitefly dsRNA bacterial solution at a volume ratio of 1:1 to avoid the whitefly dsRNA bacterial solution being degraded by ultraviolet light.

[0220] Example 34

[0221] The biological control method for whiteflies based on 5% UV protectant SH consists of the following steps:

[0222] (1) Using 0.1% (volume fraction) Tween80 aqueous solution as solvent, sodium humate was prepared into a 5% (mass fraction) solution as a 1% UV protectant SH solution.

[0223] (2) Mix 5% UV protectant SH solution with a solution of dsGawky cells (i.e., cells that transcribe the whitefly Gawky gene dsRNA) (concentration 2 g / L) at a volume ratio of 1:1. After shaking and mixing, a mixture of 0.2% UV protectant SH and dsGawky cells for whitefly RNAi interference is obtained.

[0224] (3) The above-mentioned 5% UV protectant SH and the transcribed dsGawky bacteria were sprayed on the plants, causing the whiteflies to feed on the sprayed plants, thereby increasing the mortality rate of the whiteflies.

[0225] The method for preparing the solution for transcribing dsGawky cells is the same as in Example 3.

[0226] Example 35

[0227] The lethal effect of indoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 5% UV protectant SH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0228] The transcribed dsGawky bacterial cell solution prepared in Example 34 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator (no sunlight) at 27 ± 1°C, relative humidity 60% ± 5%, and a photoperiod of 16 L:8 D. Adult mortality was recorded 7 days after spraying.

[0229] The mixture of 5% UV protectant SH prepared in Example 34 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed in a constant temperature incubator with no sunlight, a temperature of 27±1℃, a relative humidity of 60%±5%, and a photoperiod of 16L:8D. Adult mortality was recorded 7 days after spraying.

[0230] The results are as follows Figure 21 As shown, under indoor conditions, the mortality rate of adult Q-type whiteflies was 53.9% when sprayed with a solution of transcribed dsGawky bacteria, and 52.6% when sprayed with a mixture of 5% UV protectant SH and transcribed dsGawky bacteria.

[0231] Example 36

[0232] The lethal effect of outdoor spraying of bacterial solution transcribed from dsGawky bacteria or a mixture of 5% UV protectant SH and transcribed from dsGawky bacteria on adult Q-type whiteflies.

[0233] The transcribed dsGawky bacterial cell solution prepared in Example 34 was dissolved and diluted with ddH2O to a wet bacterial solution concentration of 2 g / L. This solution was then sprayed onto cotton plants, three times on each side of the leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates per treatment. The plants were placed outdoors in sunlight. Adult mortality rates were recorded 7 days after spraying.

[0234] The mixture of 5% UV protectant SH prepared in Example 34 and transcribed dsGawky bacteria was sprayed onto cotton plants, three times on each side of the cotton leaves. After the solution dried, 30 adult Q-type whiteflies were released onto the cotton plants, with three replicates for each treatment. The plants were placed outdoors in sunlight. The adult mortality rate was recorded 7 days after spraying.

[0235] The results are as follows Figure 22 As shown, under outdoor conditions, the mortality rate of adult Q-type whiteflies was 18% when sprayed with a solution of transcribed dsGawky bacteria, while the mortality rate of adult Q-type whiteflies was 64.8% when sprayed with a mixture of 5% UV protectant SH and transcribed dsGawky bacteria. The two results showed a significant difference.

[0236] Different protective agents were sprayed on cotton under outdoor conditions with a mixture of transcribed dsGawky bacteria. The lethality of the Q-type whitefly adults was observed after 7 days: the control efficacy of 0.2% UV protectant KH and 0.2% UV protectant SH did not meet expectations. Although the control efficacy of 5% UV protectant KH and 5% UV protectant SH was better than that of 1% UV protectant KH and 1% UV protectant SH, the material cost was higher. Considering cost and other factors, 1% UV protectant KH and 1% UV protectant SH were selected.

[0237] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention in any other way. Any person skilled in the art may make changes or modifications to the above-disclosed technical content to create equivalent embodiments. However, any simple modifications, equivalent changes, and modifications made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention shall still fall within the protection scope of the present invention.

Claims

1. A method for avoiding UV degradation of a dsRNA bacterial solution, comprising, The dsRNA bacterial solution is mixed with a UV protective agent to prepare a mixture of the UV protective agent and the dsRNA bacterial solution; the UV protective agent is one of potassium humate and sodium humate; the specific method is as follows: (1) The UV protective agent is dispersed in a 0.1% Tween 80 aqueous solution to prepare a 1% UV protective agent solution; (2) The UV protective agent solution is mixed with a bacterial solution for transcribing dsRNA at a volume ratio of 1:1, and the mixture is shaken and mixed uniformly; the bacteria are Escherichia coli.

2. The method for avoiding the degradation of a dsRNA bacterial solution by ultraviolet light according to claim 1 is applied to the prevention and control of agricultural pests by RNA interference.

3. A biological preparation for controlling Bemisia tabaci, characterized by, The dsRNA bacterial solution for interfering with the Gawky gene of Bemisia tabaci comprises a UV protective agent and a dsRNA bacterial solution for interfering with the Gawky gene of Bemisia tabaci; the UV protective agent is one of potassium humate and sodium humate; The dsRNA bacterial solution for interfering with the Gawky gene of Bemisia tabaci is a bacterial solution for transcribing the dsRNA of the Gawky gene of Bemisia tabaci; The preparation method of the biological agent for preventing and controlling Bemisia tabaci is as follows: (1) The UV protective agent is dispersed in a 0.1% Tween 80 aqueous solution to prepare a 1% UV protective agent solution; (2) The UV protective agent solution is mixed with a bacterial solution for transcribing the dsRNA of the Gawky gene of Bemisia tabaci at a volume ratio of 1:1, and the mixture is shaken and mixed uniformly; the bacteria are Escherichia coli, thereby obtaining the biological agent for preventing and controlling Bemisia tabaci; The bacterial solution for transcribing the dsRNA of the Gawky gene of Bemisia tabaci is prepared by the following method: The dsRNA sequence of the Gawky gene is amplified by using primers, and the amplified sequence is connected to an expression vector to construct a recombinant vector, which is transformed into an expression strain; the expression strain is induced to transcribe the dsRNA sequence of the Gawky gene, the bacterial solution is diluted with ddH2O after the supernatant is discarded, thereby obtaining the bacterial solution; The nucleic acid sequence of the Gawky gene is shown in SEQ ID NO: 1; The primers for amplifying the dsRNA sequence of the Gawky gene are shown in SEQ ID NO: 2 and SEQ ID NO:

3.

4. The biological preparation for controlling Bemisia tabaci according to claim 3, wherein, The method for inducing the expression strain to transcribe the dsRNA sequence of the Gawky gene is as follows: The recombinant bacteria transformed with the recombinant vector expressing dsRNA of the Bemisia tabaci Gawky gene were inoculated into LB medium containing Amp and cultured at 37°C with shaking at 20 r / min. When the OD value of the bacterial solution reached 0.5, an IPTG solution was added to a final concentration of 0.5 M for induction for 4 h, and the product was obtained. 600 The recombinant bacteria transformed with the recombinant vector expressing dsRNA of the Bemisia tabaci Gawky gene were inoculated into LB medium containing Amp and cultured at 37°C with shaking at 20 r / min. When the OD value of the bacterial solution reached 0.5, an IPTG solution was added to a final concentration of 0.5 M for induction for 4 h, and the product was obtained 5. The biological agent for preventing and controlling Bemisia tabaci according to claim 4 is applied to the biological prevention and control of Bemisia tabaci.

6. A method for biological control of Bemisia tabaci, characterized by, The biological agent for preventing and controlling Bemisia tabaci according to claim 4 is sprayed on plants, so that Bemisia tabaci feeds on the sprayed plants, thereby increasing the mortality rate of Bemisia tabaci.

7. The method according to claim 6, wherein the method is characterized by, The Bemisia tabaci is Q-type Bemisia tabaci or B-type Bemisia tabaci.

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