Application of CmLBD29 gene in improvement of grafting affinity of cucurbitaceae plants

Abstract

The invention relates to the technical field of biology, in particular to application of a CmLBD29 gene to improvement of grafting affinity of cucurbitaceae plants. The invention discloses an application of a CmLBD29 gene or an encoded protein thereof in improvement of grafting affinity of cucurbitaceae plants. According to the grafted plant with the silent CmLBD29 gene, the incidence rate of adventitious roots at the joint is remarkably reduced, and the healing quality is improved. Meanwhile, after the CmLBD29 gene is silenced, the scion plant height, stem diameter, dry weight, fresh weight and the SPAD value of scion leaves of the grafted plant are all remarkably improved, multi-dimensional collaborative optimization of the growth vigor and resource utilization efficiency of the grafted plant is completed, a brand new molecular regulation target is provided for improvement of grafting affinity of cucurbitaceae crops, and the method has a wide application prospect. The method has important application value in the field of stress-resistant high-yield grafted germplasm creation.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of biotechnology, and particularly relates to application of CmLBD29 gene in improving graft compatibility of Cucurbitaceae plants. BACKGROUND

[0002] As a core agronomic means to cope with continuous cropping obstacles and enhance stress resistance of Cucurbitaceae crops, grafting technology is widely used to resist soil-borne diseases such as Fusarium wilt and root-knot nematode, so as to improve the yield and environmental adaptability of crops. However, this technology system is plagued by two major defects in practice: on the one hand, Cucumis melo and Lagenaria siceraria grafting shows symbiotic incompatibility effect, which is manifested as inhibition of plant height and stem diameter of grafted plants, and wilting or even death of grafted seedlings about 35 days after grafting. On the other hand, a large number of adventitious roots appear in the region 2 cm above the grafting interface of Cucumis melo and Lagenaria siceraria, which not only consumes photosynthetic products, but also seriously hinders the formation of vascular bridge between stock and scion, and reduces the nutrient transport efficiency, becoming a key bottleneck affecting the graft compatibility of Cucumis melo and Lagenaria siceraria.

[0003] At the molecular mechanism level, LBD29 gene as a member of plant LBD (Lateral Organ Boundaries Domain) transcription factor family, the current research reports on its function analysis are focused on the internal metabolic process of single plant. It is confirmed that the gene directly activates the expression network of downstream genes such as AUX / IAA by responding to the gradient of auxin signal, and drives the formation of lateral root primordia; at the same time, it participates in the nitrogen stress response pathway, and regulates the expression level of nitrate transporter gene NRT2.1 by interacting with ARF7 / ARF19 transcription factors. However, all the above research system conclusions are derived from single plant experimental system without grafting operation. There is a significant gap in the current field in recognizing the biological function of the gene at the interface of stock-scion interaction: how it promotes the connection of transport tissues (such as xylem and phloem) between scion and stock by inhibiting the development of lateral roots during the healing process of grafted plants, so as to realize the transport of water and nutrients, make the scion grow normally on the stock, and improve the graft compatibility of grafted plants, such as significant increase in plant height, stem diameter and SPAD value of grafted plants, and other key physiological events are still unknown, which directly hinders the development of precise molecular breeding strategies targeting this target. SUMMARY

[0004] Therefore, the present application provides application of CmLBD29 gene in improving graft compatibility of Cucurbitaceae plants.

[0005] The technical scheme of the present application is implemented as follows:

[0006] In a first aspect, the present application provides an application of a CmLBD29 gene or a protein encoded by the CmLBD29 gene in improving graft compatibility of a Cucurbitaceae plant, wherein a nucleotide sequence of the CmLBD29 gene is shown as SEQ ID NO: 1, and an amino acid sequence of the protein encoded by the CmLBD29 gene is shown as SEQ ID NO: 2.

[0007] Further, in some specific embodiments, the Cucurbitaceae plant is Cucumis melo and Cucurbita pepo.

[0008] Further, in some specific embodiments, the application is an application of inhibiting expression of the CmLBD29 gene or the protein encoded by the CmLBD29 gene in improving graft compatibility of Cucumis melo and Cucurbita pepo.

[0009] In a second aspect, the present application provides an application of a biological material for inhibiting expression of a CmLBD29 gene or a protein encoded by the CmLBD29 gene in improving graft compatibility of Cucumis melo and Cucurbita pepo, wherein a nucleotide sequence of the CmLBD29 gene is shown as SEQ ID NO: 1.

[0010] Further, in some specific embodiments, the biological material is a biological material for silencing, knocking out or knocking down the CmLBD29 gene.

[0011] Further, in some specific embodiments, the biological material comprises a CmLBD29 gene VIGS silencing vector and / or an Agrobacterium containing the vector.

[0012] In a third aspect, the present application provides a method for improving graft compatibility of Cucumis melo and Cucurbita pepo, comprising the following steps:

[0013] S1, extracting Cucumis melo RNA and reverse transcribing the Cucumis melo RNA into cDNA, using the cDNA as a template and using SEQ ID NO: 4-5 as primers to perform PCR amplification to obtain a target fragment with homologous arms, wherein a sequence of the target fragment is shown as SEQ ID NO: 3;

[0014] S2, performing homologous recombination on the target fragment with homologous arms and an enzyme-digested VIGS virus vector, using a recombination product to transform E. coli competent cells, selecting a positive clone to extract a plasmid, and obtaining a recombination vector;

[0015] S3, using the recombination vector to infect Cucumis melo seeds through Agrobacterium mediation, and then using a CmLBD29 gene silencing Cucumis melo plant obtained through screening as a scion to be grafted onto a Cucurbita pepo stock to obtain a grafted plant with improved graft compatibility.

[0016] Further, in some specific embodiments, in the step S2, the VIGS virus vector is pV190.

[0017] Further, in some specific embodiments, the step S3 of infecting the melon seeds with the recombinant vector mediated by Agrobacterium includes:

[0018] S3-1, the recombinant vector is a pV190-CmLBD29 plasmid vector, GV3101 Agrobacterium competent cells are transformed with pV190-CmLBD29, positive monoclonal cells are screened, and Agrobacterium containing the pV190-CmLBD29 vector is obtained.

[0019] S3-2, the OD600 value of the Agrobacterium containing the pV190-CmLBD29 vector is adjusted to 0.3-0.5, dark treatment is performed for 1-3h, then the 1-1.5cm sprouted melon seeds are infected in the dark environment for 24-36h.

[0020] The beneficial effects of the present application at least include the following:

[0021] The present application provides application of CmLBD29 gene in improving graft compatibility of cucurbitaceae crops, successfully breaks the height inhibition bottleneck in melon and gourd grafting system, significantly reduces the occurrence rate of adventitious roots at the grafting interface (P<0.001), and effectively improves the interface healing quality. Experimental data shows that the scion plants using CmLBD29 gene silencing technology show significant growth in plant height (P<0.01) and stem diameter (P<0.01), and the dry weight (P<0.001) and fresh weight (P<0.001) accumulation capacity of the scion are obviously improved. In addition, the SPAD value (relative chlorophyll content) of the scion leaves also increases significantly (P<0.001), indicating that the photosynthetic efficiency is effectively improved.

[0022] The present application provides a new molecular regulation target for improving graft compatibility of cucurbitaceae crops, and realizes multi-dimensional optimization of growth potential, interface healing and resource utilization efficiency of grafted plants by precisely silencing CmLBD29 gene expression. The present application not only lays a foundation for solving the problem of incompatibility of cucurbitaceae crops grafting, but also creates a new strategy for improving stock-scion interaction through gene silencing technology, and has important application value in the field of creating stress-resistant and high-yield grafted varieties.

[0023] TERMS

[0024] In the present application, the term "VIGS" (Virus-Induced Gene Silencing) is a technology of inserting a target gene fragment (200-350bp) into a virus vector to construct a recombinant virus, and after inoculation, the plant recognizes the viral RNA as an exogenous invader, triggering the RNA silencing mechanism, i.e. DCL protein cutting viral double-stranded RNA to generate 21-24nt siRNA, siRNA combining with AGO protein to form RISC complex, degrading homologous mRNA through base complementation, and finally specifically silencing the target gene.

[0025] The term "graft compatibility" is a comprehensive biological index of the ability of the stock and the scion to form a symbiotic system through grafting, tissue healing, physiological integration, and long-term synergistic growth. BRIEF DESCRIPTION OF DRAWINGS

[0026] In order to more clearly illustrate the technical solutions in the embodiments of the present application, the drawings needed in the embodiment description will be briefly introduced. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can be obtained by those skilled in the art without creative labor.

[0027] Figure 1 It is a schematic diagram of the pV190-CmLBD29 vector of the present application;

[0028] Figure 2 It is the result of qRT-PCR detection of the relative expression amount of CmLBD29 gene, wherein "NI" is a blank control plant not immersed with Agrobacterium, "pV190" is a melon plant immersed with Agrobacterium containing pV190 empty plasmid, and "pV190-CmLBD29" is a melon plant immersed with Agrobacterium containing CmLBD29 gene silencing vector (the same below);

[0029] Figure 3 It is the number of adventitious roots of melon scions grafted with cucurbit on the 5th day and the 7th day after treatment, wherein ** indicates P<0.01, and *** indicates P<0.001 (the same below);

[0030] Figure 4 It is the plant height of melon scions grafted with cucurbit on the 21st day after treatment;

[0031] Figure 5 It is the stem diameter of melon scions grafted with cucurbit on the 21st day after treatment;

[0032] Figure 6 It is the fresh weight of melon scions grafted with cucurbit on the 21st day after treatment;

[0033] Figure 7The dry weight of the grafted melon scion on the 21st day after grafting with Cucurbita moschata;

[0034] Figure 8 The SPAD value of the grafted melon scion on the 21st day after grafting with Cucurbita moschata;

[0035] Figure 9 The plant growth of the grafted melon scion on the 21st day after grafting with Cucurbita moschata. DETAILED DESCRIPTION

[0036] In order to make the objects, technical solutions and advantages of the present application clearer, the technical solutions in the present application will be clearly and completely described below. Obviously, the described embodiments are part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor fall within the protection scope of the present application. If the specific conditions are not mentioned in the embodiments, the conventional conditions or the conditions recommended by the manufacturer are adopted. If the manufacturers of the reagents or instruments are not mentioned, they are all conventional products that can be purchased in the market. If the schemes and formulas that are not described in detail in the embodiments are not described in detail, please refer to Molecular Cloning Experiments Guide, Third Edition, Science Press.

[0037] Table 1 Sequence information table

[0038]

[0039]

[0040] EMBODIMENT

[0041] 1. Main material sources

[0042] The pV190 plasmid was a gift from Professor Guqinsheng of the Zhengzhou Fruit Tree Research Institute of the Chinese Academy of Agricultural Sciences. The plasmid vector has been published in Acucumber green mottle mosaic virus vector for virus-induced gene silencing in cucurbit plants (Liu M, Liang Z, Aranda MA, et al. Plant Methods. 2020; 16(1): 9).

[0043] The E. coli competent DH5a and Agrobacterium GV3101 were purchased from Shanghai Uptide Biotechnology Co., Ltd.

[0044] TransZol RNA Extraction Kit (purchased from Beijing Zoman Biotechnology Co., Ltd.); ToloScript All-in-one RT EasyMix for qPCR (purchased from Shanghai Tuoluogang Biotechnology Co., Ltd.); 2x Super Pfx MasterMix (purchased from Nanjing Novozyme Biotech Co., Ltd.); QIAquick PCR Purification Kit (purchased from QIAGEN, Germany); PerfectStart Green qPCR SuperMix (purchased from Beijing Zoman Biotechnology Co., Ltd.).

[0045] The primers used in the embodiments of the present application were all custom synthesized by Wuhan Hechong Gene Technology Co., Ltd.

[0046] 2. Preparation of VIGS silencing vector of CmLBD29 gene (pV190-CmLBD29 plasmid)

[0047] (1) Selecting the target sequence for silencing CmLBD29 gene

[0048] The CDS sequence of CmLBD29 gene obtained from CuGenDB (MELO3C012908 (gene) Melon (DHL92) v3.5.1) database is shown as SEQ ID NO: 1; according to the selection principle of silencing fragment in VIGS technology, a specific silencing fragment of CmLBD29 gene is designed, and the sequence is shown as SEQ ID NO: 3.

[0049] (2) PCR amplification of CmLBD29 gene silencing fragment

[0050] Total RNA was extracted from the hypocotyls of melon stem segments, and cDNA was obtained by reverse transcription, and then CmLBD29-VIGS-F and CmLBD29-VIGS-R (shown as SEQ ID NO: 4-5, respectively) were used as primer pairs to amplify the silencing fragment by PCR, and the amplification product was electrophoresed, and the cDNA of the CmLBD29 gene silencing fragment was recovered from the correct electrophoresis band. The specific operation is as follows:

[0051] A. Extract total RNA from the hypocotyls of melon stem segments:

[0052] The hypocotyl of melon stem section was ground into powder with liquid nitrogen, and then TransZol reagent was added for homogenate treatment. The mixing ratio was 100 mg of ground powder to 1 mL of TransZol reagent. After treatment by a homogenizer, the mixture was mixed by repeatedly blowing and sucking with a pipette, and then incubated at room temperature for 5 minutes. Then, RNA extraction reagent (0.2 mL of RNA Extraction Agent for every 1 mL of TransZol) was added to the mixture, which was incubated at room temperature for 3 minutes after being vigorously shaken for 15 seconds. The mixture was centrifuged at 10,000 rpm at 4°C for 15 minutes, and the solution was separated into a colorless aqueous phase (upper layer), an intermediate layer, and a pink organic phase (lower layer). RNA was mainly enriched in the aqueous phase, which accounted for about 60% of the initial volume of TransZol. The aqueous phase was carefully transferred to a new centrifuge tube, and isopropanol (0.5 mL of isopropanol for every 1 mL of TransZol) was added. After mixing by inversion and incubation at room temperature for 10 minutes, the mixture was centrifuged at 10,000 rpm at 4°C for 10 minutes. After the supernatant was discarded, a gelatinous precipitate was formed on the side and bottom of the tube. 1 mL of 75% (v / v) ethanol (prepared by DEPC-treated water) was added to the precipitate, which was mixed by vigorous vortexing and then centrifuged at 7,500 rpm at 4°C for 5 minutes. The supernatant was discarded (to remove salt ions in the RNA, the ethanol should be removed as much as possible), and the precipitate was air-dried at room temperature for about 5 minutes. The precipitate was dissolved in 50 μL of RNA dissolving solution, and incubated at 60°C for 10 minutes. The final sample was stored at -70°C for use, which was a total RNA-containing solution

[0053] B, reverse transcription to obtain cDNA

[0054] A 20 μL reverse transcription reaction system was constructed, and the components were as follows:

[0055] COMPOSITIONS VOLUME RNA template 2 μL 5x All-in-one RT Buffer 4 μL All-in-one Enzyme Mix 1 μL RNase-free ddH2O 13 μL

[0056] The PCR instrument was used for reaction at 50°C for 15 minutes, and then at 85°C for 5 seconds to obtain the cDNA template.

[0057] C, PCR amplification of CmLBD29 gene silencing fragment

[0058] A 50 μL PCR reaction system was constructed, and the components were as follows:

[0059]

[0060]

[0061] The PCR reaction conditions were as follows:

[0062]

[0063] The product with homologous arms was finally obtained. After agarose gel electrophoresis detection, the gel was recovered to obtain a DNA fragment with a size of about 300 bp, and the concentration was 86 ng / μL.

[0064] (3) Construction of pV190-CmLBD29 plasmid

[0065] The pV190 plasmid was single-digested, and the enzyme digestion reaction system (total 20 μL) was as follows: 1 μL pV190 plasmid, 1 μL BamHI, 2 μL 10×KBuffer, and the rest was double distilled water. Incubation at 37°C for 3 h.

[0066] The digested pV190 fragment and the PCR reaction product (300 bp DNA fragment) of the previous step were homologously recombined, and the reaction system (total 10 μL) was as follows: 2 μL 5×In-Fusion HD Enzyme Premix, 50 ng digested pV190 fragment, PCR reaction product of the previous step, and the rest was double distilled water. Incubation at 50°C for 15 min. Then the homologous recombination product was transferred into E. coli, and positive clones were screened from the transformants. The pV190-CmLBD29 plasmid (i.e. VIGS silencing vector of CmLBD29 gene) was obtained by extracting the culture of the positive clones. The specific operation was as follows:

[0067] 10 μL of the homologous recombination reaction product was taken into 100 μL of E. coli competent DH5α, and mixed by flicking. Incubation in ice water bath for 25 min, heat shock in 42°C water bath for 2 min, and then immediately incubate in ice water bath for 3 min. Add 1 mL of LB solution and mix well; incubate at 37°C, 200 rpm on a shaker for 30 min. About 200 μL of the transformed bacterial solution was uniformly coated on the LB solid plate containing Kanamycin (50 μg / mL), and the plate was inverted and cultured in a 37°C incubator for 12 h.

[0068] The positive single colony was picked and inoculated into LB liquid medium containing Kanamycin (50 μg / mL), and incubated at 37°C, 220 rpm on a shaker for 12 h. 1 mL of the bacterial solution was used for colony PCR verification, and the PCR product was detected by electrophoresis. The positive clone bacterial solution was sent to Wuhan Hechuan Biotechnology Company for sequencing and sequence alignment with NCBI. If the alignment is correct, it means that the VIGS silencing vector is successfully constructed. The plasmid was extracted from the verified positive bacterial solution to obtain the VIGS silencing vector of CmLBD29 gene (i.e. pV190-CmLBD29 plasmid).

[0069] 3. Transformation of Agrobacterium GV3101

[0070] Take 10 μL pV190-CmLBD29 plasmid vector, add 100 μL of GV3101 Agrobacterium competent cells, mix gently, ice bath for 5 min, freeze in liquid nitrogen for 5 min, then quickly placed in a 37°C water bath for 5 min, ice bath for 5 min. Add 700 μL LB medium, incubate at 28°C, 220 rpm on a shaker for 3 h. Centrifuge at 12000 rpm for 1 min to collect the bacterial solution, discard the supernatant, add 100 μL LB liquid medium, resuspend, mix well, and evenly spread on LB+Kan (50 μg / mL)+Rif (50 μg / mL) plates, 28°C dark culture for 2 d. Pick out Agrobacterium monoclonal bacteria spot with LB+Kan (50 μg / mL)+Rif (50 μg / mL) liquid medium, shake bacteria for 12 h, and obtain Agrobacterium containing pV190-CmLBD29 vector. Store the bacterial solution at -80°C ultra-low temperature refrigerator for future use.

[0071] 4、Agrobacterium infection of melon

[0072] Soak melon seeds in 60°C warm water for 1 day, and then germinate the soaked melon seeds at 28°C in the dark for 24 h. Agrobacterium containing pV190-CmLBD29 vector is cultured in LB+Kan (50 μg / mL)+Rif (50 μg / mL) liquid medium for 18 h, and the Agrobacterium bacterial solution is collected. Centrifuge the Agrobacterium bacterial solution at 6000 rpm for 8 min, and resuspend the precipitate with MAA resuspension solution (MgCl21M 5 mL; MES 0.5M 10 mL; Acetyl-syringone 20 mg / mL 200 μL and water 500 / mL). Adjust the OD value to 0.4, and treat at room temperature in the dark for 2 h. 600 600

[0073] Lay the melon seeds with about 1 cm sprouts in a 9 cm culture dish. Pour 30 mL of the above inoculum (OD 600 = 0.4) into the culture dish, which just covers the sprout but does not completely immerse the radicle, and incubate at 22°C in the dark for 24 h. Then place the inoculated seeds on sterile filter paper to absorb the excess bacterial solution, and then sow. After 14 d, if the melon leaves show light bleaching, it is a positive clone

[0074] 5、qRT-PCR verification of CmLBD29 expression

[0075] Extract melon RNA and reverse transcribe to cDNA (method as above), use qPCR-CmLBD29-F and qPCR-CmLBD29-R as primers for PCR amplification of cDNA, use CmADP gene as internal reference (primers CmADP-F and CmADP-R), use 2 -ΔΔCtThe relative expression amount of CmLBD29 in the positive clone, the melon plant not infiltrated with Agrobacterium (blank control: NI control group), and the melon plant infiltrated with Agrobacterium containing pV190 empty plasmid was detected.

[0076] A 10 μL PCR reaction system was constructed, and the components were as follows:

[0077]

[0078]

[0079] The PCR reaction conditions were as follows:

[0080]

[0081] The results are shown in Table 1. Figure 2 The relative expression amount of CmLBD29 gene of the positive clone was significantly lower than that of the NI control group and the melon plant infiltrated with Agrobacterium containing pV190 empty plasmid (P<0.001), indicating that the CmLBD29 gene silencing was successful.

[0082] 6. Grafting

[0083] The branches and leaves of the positive clone melon were grafted onto the cucurbit (one leaf and one heart stage) rootstock, covered and kept moist, and the phenotype was observed. The melon plants not infiltrated (NI group) and the melon plants infiltrated with Agrobacterium containing pV190 empty plasmid (pV190 group) were used as controls, with 15 plants in each group and 3 repetitions. The scion of the grafted plant was phenotypically identified, the number of adventitious roots was observed and recorded, and the plant height, stem diameter, dry / fresh weight, and SPAD value (SPAD-502 chlorophyll meter) were measured at 21 days after grafting. Each plant was measured 5 times, and the average value was taken. The different treatment groups were compared by t-test, and the specific index determination method was as follows:

[0084] Plant height: The height of the scion (from the healing part to the growth point of the scion) was measured with a tape measure.

[0085] Stem diameter of scion: The stem diameter of the scion, i.e. the stem diameter at the position 1-2 cm above the healing part, was measured with a vernier caliper.

[0086] Fresh weight: The scion was cut at the position of the grafting interface 21 days after grafting, and the weight was measured with a balance and labeled.

[0087] Dry weight: The fresh scion was placed in an oven at 80°C for drying for 3 days, and the weight was measured with a balance and labeled to correspond to the fresh weight.

[0088] SPAD value: The SPAD value (chlorophyll content) of the second fully expanded true leaf from top to bottom.

[0089] Results:

[0090] As shown in Table 2, the plant height of the pV190 group was significantly lower than that of the NI group (P<0.001), indicating that the CmLBD29 gene silencing affected the plant height of the melon plant.Figure 3 As shown, on days 5 and 7 post-grafting, compared with the control, the number of adventitious roots in CmLBD29 gene-silenced plants was significantly reduced (P < 0.001). The reduction in adventitious root number after CmLBD29 gene silencing indicates that the CmLBD29 gene promotes adventitious root formation. This reduction in adventitious root number avoids resource competition with the vascular bundles at the grafting interface, promotes direct communication between the vascular bundles of the rootstock and scion, and thus improves the grafting interface healing efficiency.

[0091] like Figure 4 and Figure 5 As shown, on the 21st day after grafting, the height of the scion in the CmLBD29 gene-silenced plant was ( Figure 9 Both stem diameter and stem diameter were significantly higher than the control group (P<0.01); Figure 6 and Figure 7 As shown, the dry and fresh weights of the scions also increased significantly (P<0.001), demonstrating improved vascular system transport efficiency. Figure 8 As shown, the SPAD value of plants with silenced CmLBD29 gene was significantly increased (P<0.001), indicating enhanced photosynthetic efficiency.

[0092] The above results indicate that silencing the CmLBD29 gene can significantly improve the grafting compatibility between melon (scion) and gourd (rootstock), verifying that this gene, as a key target, can improve the growth vigor and graft union healing quality of grafted crops, enhance grafting compatibility, and improve the overall vigor and resource utilization efficiency of the graft, providing a new strategy for creating stress-resistant and high-yield grafting inoculum.

[0093] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. The application of CmLBD29 gene or its coded protein in improving the graft compatibility of Cucurbitaceae plants, characterized in that, The nucleotide sequence of the CmLBD29 gene is shown as SEQ ID NO:

1.

2. Use according to claim 1, characterized in that, The Cucurbitaceae plants are melon and calabash.

3. Use according to claim 2, characterized in that, The application of inhibiting the expression of CmLBD29 gene or its coded protein in improving the grafting affinity of melon and calabash.

4. The use of biological material inhibiting the expression of CmLBD29 gene or its coded protein in improving the compatibility of melon and gourd grafting, characterized in that, The nucleotide sequence of the CmLBD29 gene is shown as SEQ ID NO:

1.

5. Use according to claim 4, characterized in that, The biological material is a biological material for silencing, knocking out or knocking down the CmLBD29 gene.

6. Use according to claim 4, characterized in that, The biological material comprises a CmLBD29 gene VIGS silencing vector and / or agrobacterium containing the vector.

7. A method of improving the compatibility of melon and cucurbit grafting, characterized by, The method comprises the following steps: S1, extracting melon RNA and reverse transcribing into cDNA, taking the cDNA as a template and taking SEQ ID NO: 4-5 as primers to perform PCR amplification to obtain a target fragment with homologous arms, and the sequence of the target fragment is shown as SEQ ID NO: 3; S2, performing homologous recombination on the target fragment with homologous arms and the enzyme-digested VIGS virus vector, transforming E. coli competent cells with the recombination product, extracting plasmids from positive clones to obtain a recombination vector; S3, infecting melon seeds with the recombination vector through agrobacterium mediation, then grafting the CmLBD29 gene silenced melon plants obtained through screening to the calabash stock to obtain a grafted plant with improved grafting affinity.

8. The method of claim 7, wherein, In the step S2, the VIGS virus vector is pV190.

9. The method of claim 8, wherein, In the step S3, the step of infecting melon seeds with the recombination vector through agrobacterium mediation comprises: S3-1, the recombination vector is a pV190-CmLBD29 plasmid vector, the pV190-CmLBD29 is used to transform GV3101 agrobacterium competent cells, positive monoclonal is screened, and agrobacterium containing the pV190-CmLBD29 vector is obtained. S3-2, OD of Agrobacterium tumefaciens bacterial suspension containing pV190-CmLBD29 vector 600 Adjust the value to 0.3-0.5, dark treat for 1-3 hours, and then infect melon seeds that have sprouted 1-1.5cm in the dark for 24-36 hours.

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