Ljcyd2 gene and application thereof in regulating plant leaf trichome density

Abstract

The application discloses an LjCYCD2 gene and application thereof in regulating plant leaf trichome density, and belongs to the technical field of genetic engineering.The amino acid sequence coded by the LjCYCD2 gene is shown as SEQ ID NO:2, and the nucleic acid sequence thereof is shown as SEQ ID NO:1.The gene is expressed in Lonicera japonica leaves, and the expression amount gradually decreases with the maturity of the leaves;after the LjCYCD2 gene of the Lonicera japonica leaves is silenced by VIGS, the trichome density of the Lonicera japonica leaves is obviously reduced.A plant overexpression vector of the LjCYCD2 gene of the Lonicera japonica is constructed, and is transformed into tobacco, and the obtained transgenic tobacco leaves have an obviously increased number of trichomes.It can be seen that the LjCYCD2 gene of the Lonicera japonica is related to the development of plant leaf trichomes, and has important application value in regulating the trichome density on the surface of plant leaves.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of genetic engineering, and particularly relates to LjCYCD2 gene and application thereof in regulating plant leaf trichome density. BACKGROUND

[0002] Trichomes are usually distributed on the surface of plant stems, leaves, flowers, fruits and other organs, and can not only perform the function of physical obstruction like non-trichomes, but also can secrete terpenes, alkaloids, phenols and sterols and other secondary metabolites. Studies have shown that trichomes can regulate the response of plants to abiotic stress, help plants resist low temperature and improve tolerance in heavy metal contaminated soil; and can also resist animal damage, such as the trichomes of Solanaceae plants can produce acyl glycoside metabolites with insecticidal activity, and metabolites such as delta-elemene and beta-caryophyllene produced in tomato (Lycopersicon esculentum) trichomes not only improve the tolerance of tomatoes to red spider mites, but also have a repellent effect on a variety of herbivorous animals. The trichomes of many medicinal plants are also the sites where medicinal ingredients are produced. Artemisinin is a sesquiterpene lactone produced in the trichomes of Artemisia annua, which can be used as an antimalarial drug; the shield-shaped trichomes on the surface of Chinese catnip (Nepeta cataria) contain a large amount of volatile oil, which is the main medicinal ingredient.

[0003] Trichomes are a kind of exocrine structure derived from epidermal initial cells through cell division. In the early development of trichomes, certain initial cells in the epidermis receive cell division signals that can form trichomes, and then undergo cell division through strict regulation of the cell cycle. Different numbers, directions and cell extension degrees of these cell divisions will lead to the formation of different types of trichomes. The density and size of trichomes are important factors for regulating the content of metabolites, so increasing the density of trichomes is considered an effective way to increase the content of secondary metabolites.

[0004] Cell cycle regulation plays a key role in plant cell differentiation, resulting in the formation of single-cell or multi-cell trichomes. Studies have shown that cell cycle-related genes play an important role in the early stages of trichome development, but these studies have mostly focused on single-cell trichomes, such as in Arabidopsis thaliana, where SIM can effectively promote cell division and cell differentiation of the trichome body. There are few studies on the molecular regulation mechanism of multi-cell trichome development. In the study of Artemisia annua multi-cell trichomes (Artemisia annua trichomes are composed of 10 cells, which are two basal cells, two stalk cells, four lower top cells and two top cells), it was found that overexpression of TLR1 and TLR2 can significantly reduce the density of trichomes and the content of artemisinin. The CsMYB6 gene in Cucumis sativus plays an important role in regulating the development of cucumber fruit trichomes, and forms a CsMyb6-CsTRY complex to inhibit the formation of trichomes on the surface of cucumber fruit.

[0005] Lonicera japonica is a climbing shrub of Caprifoliaceae Lonicera, the flowers of which are called Jinyinhua and the stems and leaves of which are called Lonicera japonica, which are traditional Chinese medicinal materials, rich in active ingredients such as organic acids, triterpenes and flavonoids, and have good effects in treating cold, headache, fever and rash. Studying the development-related genes of Lonicera japonica trichomes can provide further basis for the development and molecular regulation of multi-cell trichomes and provide reference for the cultivation of high-quality varieties of Lonicera japonica. SUMMARY

[0006] In view of the problems in the prior art, the purpose of the present application is to provide LjCYCD2 gene and its application in regulating the density of plant leaf trichomes.

[0007] In order to achieve the above purpose, the technical scheme adopted by the present application is as follows:

[0008] LjCYCD2 gene, the amino acid sequence encoded by which is shown in SEQ ID NO: 2, and the nucleic acid sequence thereof is shown in SEQ ID NO: 1.

[0009] The application of the above-mentioned LjCYCD2 gene in regulating the density of plant leaf trichomes.

[0010] A method for regulating the density of plant leaf trichomes, by silencing the LjCYCD2 gene in the plant to reduce the density of plant leaf trichomes, or by overexpressing the LjCYCD2 gene in the plant to increase the density of plant leaf trichomes, wherein the amino acid sequence of the LjCYCD2 gene is shown in SEQ ID NO: 2.

[0011] On the basis of the above scheme, the nucleic acid sequence of the LjCYCD2 gene is shown as SEQ ID NO: 1.

[0012] On the basis of the above scheme, a silencing vector or overexpression vector containing the LjCYCD2 gene is constructed, and the plant body is transformed, so that the LjCYCD2 gene in the plant body is silenced or overexpressed, thereby reducing or increasing the leaf trichome density of the plant.

[0013] On the basis of the above scheme, the method for transforming the plant body is one of Agrobacterium-mediated method, gene gun method, electric shock method, PEG method and liposome method.

[0014] On the basis of the above scheme, the Agrobacterium is Agrobacterium tumefaciens or Agrobacterium rhizogenes.

[0015] On the basis of the above scheme, the plant is honeysuckle or tobacco.

[0016] The above LjCYCD2 gene is used for preparing a product for regulating the leaf trichome density of a plant.

[0017] A product for regulating the leaf trichome density of a plant, which is a recombinant expression vector, an expression cassette, a recombinant bacterium, a recombinant virus or a transgenic cell line containing the LjCYCD2 gene sequence; the nucleic acid sequence of the LjCYCD2 gene is shown as SEQ ID NO: 1.

[0018] Advantages of the technical scheme of the present application

[0019] The LjCYCD2 gene of honeysuckle is cloned, which is expressed in the leaves of honeysuckle, and the expression amount gradually decreases with the maturity of the leaves. After silencing the LjCYCD2 gene of honeysuckle leaves by VIGS, the trichome density of the leaves of honeysuckle is obviously reduced. The plant overexpression vector of the LjCYCD2 gene of honeysuckle is constructed, and tobacco is transformed, and the number of trichomes on the leaves of the obtained transgenic tobacco is obviously increased. It can be seen that the LjCYCD2 gene of honeysuckle is related to the development of the trichomes on the leaves of plants, and has important application value in regulating the trichome density on the surface of the leaves of plants. BRIEF DESCRIPTION OF DRAWINGS

[0020] Figure 1 The relative expression amount of the LjCYCD2 gene in different leaves of honeysuckle;

[0021] Figure 2 The relative expression level of the LjCYCD2 gene in the first pair of leaves of honeysuckle after gene silencing (error bar represents SD; asterisk represents significant difference, one-way ANOVA, *P<0.1, **P<0.01, ***P<0.001, ****P<0.0001);

[0022] Figure 3 Phenotype and trichome density of Lonicera japonica leaf after silencing LjCYCD2 gene by VIGS (A: TRV1-TRV2, B: TRV2 / LjCYCD2, C: Change of trichome density after different treatments);

[0023] Figure 4 qRT-PCR analysis of relative expression level of LjCYCD2 gene in transgenic tobacco leaf after overexpression of LjCYCD2 gene;

[0024] Figure 5 Phenotype and trichome density of transgenic tobacco leaf (A: pSuper1300, B: OE-LjCYCD2, C: Change of trichome density after different treatments). DETAILED DESCRIPTION

[0025] The terms used in the present application have the meanings generally understood by those of ordinary skill in the art, unless otherwise defined. The present application is described in further detail below in conjunction with specific examples and with reference to the data. The following examples are merely for the purpose of illustration of the present application and do not limit the scope of the present application in any way.

[0026] The experimental methods in the following examples are all conventional methods, and are carried out according to the techniques or conditions described in the literature in the art or according to the product instructions, unless otherwise specified. The experimental materials, reagents, and drugs used in the following examples can be purchased through general channels, unless otherwise specified.

[0027] In the following examples,

[0028] Lonicera japonica "Beihua No. 1": obtained from Linyi, Shandong; Nicotiana benthamiana: provided by the Plant Genetics and Development Laboratory of Qingdao Agricultural University; pSuper1300 vector and TRV2 vector: provided by the Plant Genetics and Development Laboratory of Qingdao Agricultural University; P19 and TRV1: provided by the Plant Genetics and Development Laboratory of Qingdao Agricultural University; E. coli DH5a competent cells, Agrobacterium tumefaciens EHA105, and Agrobacterium tumefaciens GV3101: Shanghai Weidi Biotechnology Co., Ltd.

[0029] Example 1

[0030] Cloning of LjCYCD2 gene

[0031] Total RNA was extracted from the leaves of Lonicera japonica "Beihua No. 1", and reverse-transcribed into cDNA as a template. The LjCYCD2 gene was amplified by PCR using specific primers, and the nucleic acid sequence of the LjCYCD2 gene obtained after sequencing is shown as SEQ ID NO: 1, and the encoded amino acid sequence is shown as SEQ ID NO: 2.

[0032] SEQ ID NO: 1 (5'→ 3')

[0033]

[0034] SEQ ID NO: 2:

[0035] MAVTPTSPSSKSISTSQNNKNKKQKPHTKIKKKNNKNKNKNDDHNPTPIINPINPQQLPPSKPVKRLKSPGVRVIGARIYDSEHGKTCHQCRQKTMDFVVSCKSQTQNKNNKQCTFRFCHKCLLNRYGEKAEELAELVDWICPKCRGICNCSFCMKKRGFHPTGILVHTAKANGFSSVSDMLLIDNSAKAETEEGSPSKKRAAKSENGDLELKRSRKDDNKEQQSRGIIKDAGASSKKATALAKEPAVGPEKAQKNVTGLDGNSDMKSQPESLKTKTNKKDTKNTNLEGLRESNEIDDSSETSMNNGAPAENITTKNDIELIKTNSEALMPNKVSTRSIKIEDNESEVPSNINVNTTAAKSRIVDEFDQFTKTEEVQGKDSVAEIPLPQGTELTNVANIDMPAEDVGHALQFLEFCEAFGKVLDLKKGQPEMLLRDLACDRRSRRKDESSVVQFHIKLLSMIQETSGKEFSSSKATVRNPWLEGLRKCISESQDPSKESLLGFLDVGVDGYEKLDTSKKLRLLTFLCDEALGTPVLRSWIDEQNSEYVKEVKKAKETILPAQKEKEKNMKRKVQNEVARAIIMNNGVPLSIAEHENLVFKIRAEAAQTLAETLEEIEMLTKTLPKETPRSDAVRSEPILFDASGRIFWKLRGYDGSSDILLQDIGSGELVTCKDRWFAYDVQEKEMVEKYLSSKRNL

[0036] The specific primer sequences of the LjCYCD2 gene are as follows:

[0037] Forward primer: 5'-atacaccaaatcgactctagaATGGCTGTAACTCCTACATCTCCTT-3' (SEQ ID NO: 3);

[0038] Reverse primer: 5'- gcccttgctcaccatggtaccTCAAAGATTCCTTTTTGATGAAAGAT-3' (SEQ ID NO: 4);

[0039] The reaction system of PCR: PrimeSTARMax DNA Polymerase (2x) 12.5 μL, Primer F / R (10 μM) 1 μL; cDNA 1 μL; RNase Free Water 9.5 μL.

[0040] The reaction procedure of PCR is as follows:

[0041]

[0042] Example 2

[0043] Expression analysis of LjCYCD2 gene

[0044] The first pair of leaves, the second pair of leaves, the third pair of leaves and the fourth pair of leaves of Lonicera japonica "Beihua No. 1" were respectively quickly frozen in liquid nitrogen and stored at -80℃. The total RNA of the above leaves was extracted according to the kit instruction of Accurate Biotechnology (Hunan) Co., Ltd., and was reversely transcribed into cDNA as a template. qRT-PCR was carried out by using the qRT-PCR specific primers of LjCYCD2 gene, and the Lonja.ACT2 / 7 gene was used as an internal reference gene.

[0045] The qRT-PCR specific primer sequences of LjCYCD2 gene are as follows:

[0046] LjCYCD2-F: 5'-TCTGAGCATGGCAAGACCTG-3' (SEQ ID NO: 5);

[0047] LjCYCD2-R: 5'-TAGGGTGAAAACCCCGCTTC-3' (SEQ ID NO: 6);

[0048] The qRT-PCR specific primer sequences of Lonja.ACT2 / 7 gene are as follows:

[0049] Lonja.ACT2 / 7-F: 5'-CCCTAAAGCCAACAGAGAGAAG-3' (SEQ ID NO: 7);

[0050] Lonja.ACT2 / 7-R: 5'-CGACCACTAGCATACAGAGAAAG-3' (SEQ ID NO: 8);

[0051] The reaction system of qRT-PCR is: DNA Polymerase (2x) 10 μL; primer F / R (10 μM) 0.8 μL; cDNA 1 μL; RNase Free Water 7.4 μL.

[0052] The reaction condition of qRT-PCR is:

[0053]

[0054] The qRT-PCR result is shown in Figure 1 It can be seen from the above table that the density of glandular hairs of honeysuckle leaves gradually decreases with the maturity of leaves, and the maturity of leaves gradually increases from the first pair of leaves to the fourth pair of leaves, and the density of glandular hairs gradually decreases. From Figure 1 It can be seen from the above table that the density of glandular hairs of honeysuckle leaves gradually decreases with the maturity of leaves, and the maturity of leaves gradually increases from the first pair of leaves to the fourth pair of leaves, and the density of glandular hairs gradually decreases. From

[0055] Example 3

[0056] Construction of LjCYCD2 gene silencing vector

[0057] According to the sequence of Xbal and Kpnl enzyme cutting sites in TRV2 vector and the requirement of homologous recombination, the primers of LjCYCD2 specific sequence were designed and synthesized by CE Design V1.04 software, and the primer sequences are as follows:

[0058] TRV2 / LjCYCD2-F: 5'-aaggttaccgaattctctagaGACGACCACAATCCAACTCCG-3' (SEQ ID NO: 9)

[0059] NO:9)

[0060] TRV2 / LjCYCD2-R: 5'-gagacgcgtgagctcggtaccTATCCAATCAACCAATTCGGC-3' (SEQ ID NO: 10)

[0061] The RNA of honeysuckle quick-frozen at 4 different leaf positions was extracted by using the kit instruction of Accurate Biotechnology (Hunan) Co., Ltd., and the cDNA was reversely transcribed by PrimerScript RT Reagent Kit, and then mixed in equal amount as a template. The specific fragment of LjCYCD2 was amplified by using the specific primers. The amplified target fragment was detected by electrophoresis, and the band with correct position and brightness was selected and purified by QIAquick Gel Extraction Kit, and then connected with the vector pTRV2 to construct the TRV2 / LjCYCD2 gene silencing vector. The product was purified by electrophoresis using a Gel DNA Extraction Mini Kit to obtain the target fragment.

[0062] The TRV2 vector was double-digested with QuickCut restriction enzyme to obtain a linearized vector.

[0063] After the target fragment and the linearized vector were added according to the requirements of the recombination reaction, the homologous recombination ligase was added, and the recombination reaction was performed at 50°C for 1h to obtain a recombinant plasmid.

[0064] The recombinant plasmid was transformed into E. coli DH5α competent cells, and positive clones were picked and detected by PCR, and then sequenced. The correct sequencing result indicated that the LjCYCD2 gene silencing vector (TRV2 / LjCYCD2) was successfully constructed.

[0065] Example 4

[0066] Effect of silencing LjCYCD2 gene on leaf trichome density of Lonicera japonica

[0067] The silencing vector TRV2 / LjCYCD2 of Example 3 was transformed into Agrobacterium tumefaciens EHA105, and TRV1 and TRV2 were separately introduced into Agrobacterium tumefaciens EHA105. The three kinds of bacterial liquid containing TRV1, TRV2 and TRV2 / LjCYCD2 recombinant plasmid were placed in a shaker, and cultured at 28°C, 200rpm, overnight. The next day, the three kinds of bacterial liquid were secondarily expanded to OD600 = 1.0-1.2. Then the bacteria were collected, and an equal volume of infection liquid was prepared (1ml MES, 40μl AS and 1ml MgCl2 were added to 100ml H2O), and the bacteria were resuspended to OD600 = 1.0-1.2. The bacterial liquid containing TRV1 plasmid and the bacterial liquid containing TRV2 plasmid were mixed in a volume ratio of 1:1, and the bacterial liquid containing TRV1 plasmid and the bacterial liquid containing TRV2 / LjCYCD2 recombinant plasmid were mixed in a volume ratio of 1:1. After mixing, the infection was performed under dark conditions for 3 hours.

[0068] The first pair of leaves of Lonicera japonica branches were selected, and high-pressure suction infection was performed using a vacuum pump. During the suction infection, the tender branches were immersed in the infection liquid, and the suction was maintained for 4-5min, 20min and 3h, and the process was repeated twice. Then, the branches were dark-treated at room temperature for 3d, and then normally cultured. The empty vector TRV2 was used as a control (CK), and the relative expression amount of LjCYCD2 gene in the first pair of leaves of Lonicera japonica was determined by qRT-PCR after VIGS transformation for 14d, and the difference in trichome density was observed.

[0069] The relative expression amount of LjCYCD2 gene in the first pair of leaves of Lonicera japonica after silencing is shown in Table 1. Figure 2As shown, the relative expression amount of the first pair of leaves LjCYCD2 gene of Lonicera japonica after silencing appeared a more obvious decrease.

[0070] After LjCYCD2 gene silencing, the density of leaf gland of Lonicera japonica changed, see Figure 3 Under the microscope, the gland of the control leaf blade main vein base was more dense Figure 3 , the gland density of the lower surface of Lonicera japonica leaf after TRV2 / LjCYCD2 silencing appeared a more obvious decrease Figure 3 , compared with the control, the gland density decreased by 2 times Figure 3 , which was consistent with the results of gene expression, further indicating that the gene was a positive regulation gene of Lonicera japonica gland development.

[0071] Example 5

[0072] Construction of LjCYCD2 gene overexpression vector

[0073] According to the LjCYCD2 gene sequence of Lonicera japonica and the Xbal, Kpnl enzyme cutting site sequence on both sides of the pSuper1300 vector and the homologous recombination requirements, the primers of LjCYCD2 specific sequence were designed and synthesized by CE Design V1.04 software, and the primer sequences were as follows:

[0074] pSuper1300-LjCYCD2-F: 5'-atacaccaaatcgactctagaATGGCTGTAACTCCTACATCTCCTT-3'(SEQ ID NO: 11);

[0075] pSuper1300-LjCYCD2-R: 5'-gcccttgctcaccatggtaccTCAAAGATTCCTTTTTGATGAAAGAT-3'(SEQ ID NO: 12);

[0076] The RNA of Lonicera japonica quick-frozen treated 4 different leaf positions was extracted using the kit instruction of Accurate Biotechnology (Hunan) Co., Ltd., and the cDNA was reversely transcribed using PrimerScriptTM RT Reagent Kit, and then mixed in equal amounts as a template, and the specific fragments of LjCYCD2 were amplified, and the amplified target fragments were electrophoretically detected, and the correct and bright bands were selected for electrophoretic product purification using Gel DNA Extraction Mini Kit kit, and the target fragments were obtained.

[0077] ​The pSuper1300 vector was double-digested with QuickCut restriction enzymes to obtain a linearized vector.

[0078] After the target fragment and the linearized vector were added according to the requirements of the recombination reaction, the homologous recombination ligase was added, and the recombination reaction was performed at 50°C for 1h to obtain a recombinant plasmid.

[0079] The recombinant plasmid was transformed into E. coli DH5α competent cells, and positive clones were picked and detected by PCR, and then sequenced. The correct sequencing indicated that the overexpression vector pSuper1300-LjCYCD2 was successfully constructed.

[0080] Example 6

[0081] Application of overexpression of LjCYCD2 gene in improving the density of plant leaf trichomes

[0082] The overexpression vector pSuper1300-LjCYCD2 of Example 5 was transformed into Agrobacterium tumefaciens GV3101, and P19 and pSuper1300 empty vector were respectively introduced into Agrobacterium tumefaciens GV3101. The three kinds of bacteria containing P19, pSuper1300 empty vector and pSuper1300-LjCYCD2 recombinant vector were activated, and the next day, the bacterial liquid was expanded to OD600 = 0.6-0.8. Then the bacteria were collected, and an equal volume of infection liquid was prepared (1ml MES, 40μL AS, 1ml MgCl2 were added to 100ml H2O), and the bacterial suspension was resuspended to OD600 = 0.6-0.8. The bacterial liquid containing P19 plasmid and the bacterial liquid containing pSuper1300 plasmid were mixed in a volume ratio of 1:1, and the bacterial liquid containing pSuper1300 plasmid and the bacterial liquid containing pSuper1300-LjCYCD2 recombinant plasmid were mixed in a volume ratio of 1:1. After mixing, the bacteria were incubated in the dark for 3 hours, and then infected.

[0083] First, the leaves of sterile tobacco seedlings were placed in MS medium for pre-culture, and then the pre-cultured tobacco leaves were completely immersed in the infection liquid, and after 8-10min of infection, they were inoculated on co-culture medium and incubated at 28°C in the dark for 2-4d. Then, differentiation culture, bud multiplication culture, rooting culture and transplantation were performed. The leaf DNA of the empty vector and the transgenic tobacco was extracted for identification, and then the RNA was extracted to detect the expression level, and the transgenic line with high expression level was identified. The empty vector pSuper1300 was used as a control (CK), and the T1 generation tobacco seeds were collected, and after T2 generation culture, the leaves of tobacco plants with consistent growth were observed for trichome density differences.

[0084] The relative expression level of LjCYCD2 gene in tobacco overexpressing LjCYCD2 gene was as follows Figure 4As shown, the relative expression of LjCYCD2 gene was obviously increased compared with the control.

[0085] After overexpression of LjCYCD2 gene, the transgenic tobacco leaves with the same growth potential were taken, and the density of trichomes was observed under a fluorescence microscope, as shown in the figure. Figure 5 Compared with the control group pSuper1300, it can be seen that the density of trichomes of LjCYCD2 overexpression strain is obviously increased. Thus, overexpression of LjCYCD2 gene in plants can increase the number of trichomes on plant leaves.

[0086] The above is only a preferred embodiment of the present application, not other forms of the present application, any skilled in the art can use the above disclosed technical content to change or modify as equivalent embodiments of equivalent changes. But any simple modification, equivalent change and modification of the above embodiments without departing from the technical solution content of the present application, according to the technical essence of the present application, still belongs to the protection scope of the technical solution of the present application.

Claims

1. LjCYCD2 A gene characterized in that, The encoded amino acid sequence is shown as SEQ ID NO:

2.

2. The method of claim 1 LjCYCD2 gene characterized in that, The nucleic acid sequence is shown as SEQ ID NO:

1.

3. The method of claim 1 or 2 LjCYCD2 The use of a gene in modulating trichome density in a plant leaf, characterized in that, by silencing a gene in a plant LjCYCD2 reduces trichome density in leaves of the plant; or by overexpressing a gene in a plant LjCYCD2 increases trichome density in leaves of the plant; the plant is tobacco or honeysuckle.

4. A method of modulating trichome density in a leaf of a plant, the method comprising, By making plants LjCYCD2 Gene silencing reduces the density of glandular hairs in plant leaves; or it reduces the number of glandular hairs in plants. LjCYCD2 Gene overexpression increases the density of glandular hairs on plant leaves; LjCYCD2 The amino acid sequence encoded by the gene is shown in SEQ ID NO:2; the plant is tobacco or honeysuckle.

5. The method for modulating trichome density of a plant leaf according to claim 4, wherein, The nucleic acid sequence of the gene is set forth in SEQ ID NO:

1. LjCYCD2 The nucleic acid sequence of the gene is set forth in SEQ ID NO:

1.

6. The method for modulating trichome density of a plant leaf according to claim 4, wherein, Construct containing the LjCYCD2 Gene silencing or overexpression vectors are used to transform plants, thereby enabling the genes described in the text to be expressed. LjCYCD2 Gene silencing or overexpression can reduce or increase the density of glandular hairs on plant leaves.

7. The method for modulating trichome density of a plant leaf according to claim 6, wherein, The transformation method of the plant body is one of Agrobacterium-mediated method, gene gun method, electric shock method, PEG method and liposome method.

8. The method for modulating trichome density of a plant leaf according to claim 7, wherein, The Agrobacterium is Agrobacterium tumefaciens or Agrobacterium rhizogenes.

9. The method of claim 1 or 2 LjCYCD2 The use of the gene in the preparation of a product for regulating the density of leaf trichomes of a plant, characterized in that, The plant is tobacco or honeysuckle.

10. A product for regulating leaf trichome density in a plant, characterized in that, For containing LjCYCD2 a recombinant expression vector, expression cassette, recombinant bacteria, recombinant virus, or transgenic cell line of the gene; the LjCYCD2 nucleic acid sequence of the gene is shown in SEQ ID NO: 1.

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