CN111575305A - Allene oxide synthetase, coding gene CsAOS and application thereof - Google Patents

Abstract

The invention belongs to the technical field of tea processing, and in particular relates to an allene oxide synthase, an encoding gene CsAOS and applications thereof. The nucleotide sequence of the allene oxide synthase gene CsAOS is shown in SEQ ID NO.1; the amino acid sequence of the protein encoded by the allene oxide synthase gene CsAOS is shown in SEQ ID NO.2. The invention provides an allene oxide synthase gene CsAOS derived from tea and related to tea aroma, and the expression difference of the allene oxide synthase gene CsAOS in different suitable black tea and green tea varieties is significant. The invention finds and proves through in vitro transient expression and in vitro oligonucleotide antisense inhibition experiments that the expression of CsAOS can significantly affect the content of volatile substances and jasmonic acids in tea leaves, which is useful for processing tea and cultivating excellent tea tree varieties. Provide new technical means.

Description

A kind of allene oxide synthase, encoding gene CsAOS and its application

technical field

The invention belongs to the technical field of tea processing, and in particular relates to an allene oxide synthase, an encoding gene CsAOS and applications thereof.

Background technique

Black tea is a kind of fully fermented tea. It is made from suitable tea tree sprouts and leaves through a series of processes such as withering, rolling (cutting), fermentation and drying. It is the main selling tea in the international tea market. consumer favorite. The polyphenols of tea leaves undergo a series of reactions such as redox during the fermentation process of black tea, so that the color of the tea leaves changes from green to red, forming the basic characteristics and aroma of "red leaf red soup", so that black tea has a unique taste.

Tea aroma refers to the unique smell of tea that can be perceived by different concentrations and proportions of odorants that affect the olfactory nerve. The aroma of tea is one of the factors that play an important role in the quality of tea. There are many factors that affect the aroma of tea, such as tea species, cultivation conditions, natural environment, processing technology, storage methods, etc. It has been detected that the volatile components in tea are as high as several Hundreds of aroma compounds, mainly aldehydes, ketones, alcohols, esters and other aroma compounds. These ingredients are combined in different proportions to form different flavors and types of tea. Although there are many kinds of aroma components in tea, their content is very small, only 0.001%-0.05% in fresh leaves (accounting for dry matter), and only 0.01%-0.03% (accounting for dry matter) in black tea.

The current research shows that the aroma substances in black tea mainly come from three aspects, including terpenes such as geraniol, compounds produced by the degradation of carotenoids, such as ionone, etc., and the oxidation of tea cell membrane lipid peroxidation. Lipid (Oxylipins) compounds. In the green leaves of most plants, monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) account for more than 80% of green leaf membrane lipids. During tea processing, these two chloroplast membrane lipids are decomposed under the action of activated phosphatase A1 (PLA1) to produce α-linolenic acid (18:3), which is processed by lipoxygenase (LOX). ) catalyzed the generation of 13S-hydroperoxylinolenic acid (13-HPOT). 13-HPOT can produce different decomposition products under different physiological states or environmental stress. On the one hand, 13-HPOT is catalyzed and decomposed into C6-C9 volatile substances by lipid hydroperoxide lyase (HPL), including volatile substances with grassy aroma such as 3-hexenal and hexanal; on the other hand, 13- HPOT is converted into oxidized phytodienoic acid (OPDA) under the catalysis of allene oxide synthase (AOS) and allene oxidative cyclase (AOC) successively, and the OPDA generated in chloroplast is transported to peroxide by transporter In the enzyme body, jasmonic acid (JA) is finally generated through oxidative phytodienoic acid reductase (OPDAReductase) and three β-oxidations. JA is then methylated by JAMT to produce methyl jasmonate. (Fig. 1: Synthetic pathway of jasmonic acids).

SUMMARY OF THE INVENTION

In view of the problems existing in the prior art, the present invention provides an allene oxide synthase, an encoding gene CsAOS and applications thereof, aiming at solving some problems in the prior art or at least alleviating a part of the problems in the prior art.

The present invention is realized in this way, an allene oxide synthase gene CsAOS, the nucleotide sequence is shown in SEQ ID NO.1, or a DNA hybridized with the DNA sequence defined by SEQ ID NO.1 and encoding the same functional protein sequence; or a DNA molecule that has more than 90% homology with the DNA sequence defined by SEQ ID NO. 1 and encodes the same functional protein.

The present invention also discloses an allene oxide synthase, the amino acid sequence is shown in SEQ ID NO.2, or the sequence shown in SEQ ID NO.2 is subjected to substitution and/or deletion of several amino acid residues and/or Or an amino acid sequence that is added and has the same protein function; or an amino acid sequence derived from the amino acid sequence shown in SEQ ID NO. 2, which has more than 98% homology and has the same protein function.

The present invention also discloses the application of the above allene oxide synthase gene CsAOS in preparation and/or use as lipoxygenase.

The invention also discloses the application of the above allene oxide synthase gene CsAOS in adjusting the content of volatile substances in tea leaves.

Further, the volatile substances in the tea leaves are heptanal, hexanal, 2-hexenal, 2,4-hexadienal, 2,4-heptadienal, 2-hexen-1-olethyl Ester, 2-ethylhexanol, benzyl alcohol, linalool, phenethyl alcohol, nonenal, nonanol, geraniol, 3-hexenyl ester hexanoic acid, 3-hexenoic acid-3-hexene At least one of ester or hexanoic acid-hexyl ester.

The invention also discloses the application of the above allene oxide synthase gene CsAOS in adjusting the content of jasmonic acid and/or salicylic acid in tea leaves.

Further, the jasmonic acid substance is at least one of jasmonic acid, jasmonic acid amino acid and methyl jasmonate.

The invention also discloses the application of the above-mentioned allene oxide synthase gene CsAOS in adjusting the content of OPDA, a prerequisite substance of jasmonic acid in tea leaves.

The invention also discloses the application of the above allene oxide synthase gene CsAOS as a marker to screen tea varieties suitable for making green tea and/or tea varieties suitable for making black tea or black tea.

The invention also discloses the application of the above allene oxide synthase gene CsAOS in the breeding of tea trees.

The invention also discloses a method for promoting the expression level of the allene oxide synthase gene CsAOS in tea leaves, which comprises at least one treatment mode of withering, rolling and fermenting the tea leaves.

To sum up, the advantages and positive effects of the present invention are:

The invention provides an allene oxide synthase gene CsAOS derived from tea and related to tea aroma, and the protein encoded by the allene oxide synthase gene CsAOS has the function of allene oxide synthase.

The present invention finds that the expression of this gene is promoted by processing withering, rolling and fermentation of black tea by detecting the expression difference of the allene oxide synthase gene CsAOS during the processing of black tea, indicating that the gene is involved in the production process of black tea.

The invention finds and proves through in vitro transient expression and in vitro oligonucleotide antisense inhibition experiments that the expression of CsAOS can significantly affect the content of volatile substances and jasmonic acids in tea leaves, which is useful for processing tea and cultivating excellent tea tree varieties. Provide new technical means.

Description of drawings

Fig. 1 is the synthetic route of jasmonic acid substances;

Fig. 2 is the expression difference of allene oxide synthase gene CsAOS in different tissues of Shuchazao according to the embodiment of the present invention; wherein, L1: one leaf; L2: two leaves; L3: three leaves; FL: flower; FR: fruit ; S: stem; R: root; B: bud;

Fig. 3 is the expression difference of allene oxide synthase gene CsAOS in the black tea processing process of the embodiment of the present invention;

Fig. 4 is the expression of CsAOS of the embodiment of the present invention in different suitable tea plant varieties;

Fig. 5 is the expression level of allene oxide synthase gene CsAOS in the in vitro oligonucleotide antisense inhibition experiment of the embodiment of the present invention;

Fig. 6 is the measurement result of jasmonic acid after 3 days of in vitro oligonucleotide antisense inhibition experiment according to the embodiment of the present invention;

Fig. 7 is the measurement result of volatile substances after 3d in vitro oligonucleotide antisense inhibition experiment according to the embodiment of the present invention;

Fig. 8 is the expression level of the in vitro transient expression allene oxide synthase gene CsAOS of the embodiment of the present invention;

FIG. 9 is the measurement result of jasmonic acids after transient expression in vitro for 3 days according to the embodiment of the present invention.

Detailed ways

In order to make the purpose, technical scheme and advantages of the present invention clearer, the present invention will be further described in detail below in conjunction with the examples. The equipment and reagents used in each embodiment and test example can be obtained from commercial channels unless otherwise specified. The specific embodiments described herein are only used to explain the present invention, and are not intended to limit the present invention.

The present invention discloses an allene oxide synthase gene CsAOS and its encoded protein and applications, as shown in the following examples. The nucleotide sequence of the allene oxide synthase gene CsAOS involved in the present invention is shown in SEQ ID NO. 1 in the sequence table, and the amino acid sequence is shown in SEQ ID NO. 2 in the sequence table.

Materials involved in the present invention:

1. Tea tree sample: Shucha morning tea tree was planted in the Agricultural Industrial Park of Anhui Agricultural University, Hefei, Luyang District, Anhui Province, and the leaf picking conditions were 25℃～28℃.

The main production process of black tea is picking, withering, rolling, fermentation and drying. Specifically: pick one bud and one leaf of the Shuchazao plant (the fresh leaf sample at this time); spread it out at room temperature for about 16 hours (the withered sample after completion); rub the tea leaves into a ball first, then lightly rub, and then lightly ( After completion, it is a rolling sample); wrap the tea leaves with gauze for 12 hours at 30°C (fermentation sample after completion), spread out every 2 hours during the fermentation period and then continue to ferment; dry at 110°C for about 15 minutes (dry sample after completion).

Tobacco samples: Nicotiana benthamiana was grown in a light culture room at a temperature of 22°C to 25°C.

2. Escherichia coli: DH5α.

3. Vector: pGEM-T Easy.

4. LB medium: Weigh 10g of NaCl, 5g of yeast extract, and 10g of tryptone, add 950mL of ultrapure water, stir to dissolve, add water to make up to 1000mL, and sterilize by high pressure steam for 15min to obtain LB liquid medium , LB solid medium is to add 15g of agar powder to LB liquid medium.

5. Ampicillin stock solution (Amp+, 50 mg/ml): Weigh 0.5 g of ampicillin Amp, dissolve it in 10 mL of sterile water, filter and sterilize it, divide into small tubes, and store at -20°C.

6. Configure in vitro oligonucleotide antisense inhibitory buffer: centrifuge the nucleotide powder (8000rpm, 5min), add 2ml of 50mMol/L sucrose solution to each tube, shake on a vortexer for a few minutes, and observe whether the powder is completely Dissolve, if no particles are seen, stop shaking to obtain in vitro oligonucleotide antisense inhibitory buffer (hereinafter referred to as buffer), and then pipette 250ul of buffer into 96-well plates for later use. .

Example 1 Differences in the expression of the allene oxide synthase gene CsAOS in different tissues of Shuchazao, different tea varieties and in the processing of black tea

1. The expression difference of allene oxide synthase gene CsAOS in different tissues of Shuchazao

The roots, stems, flowers, fruits, buds, one leaf, two leaves and three leaves of Shuchazao plants were picked respectively, and high-throughput sequencing was used to detect the expression of the allene oxide synthase gene CsAOS in each sample.

2. Differential expression of allene oxide synthase gene CsAOS during black tea processing

Take samples of black tea during the period of picking, withering, rolling, fermentation and drying, respectively, obtain cDNA according to the method described in Example 3, and perform fluorescence quantitative PCR detection. The method is as follows, and high-throughput sequencing is used for verification.

Forward primer: ACCTCCTTCTAAACCCACCAATC

Reverse primer: ACCAGGTGGCATGTTGGCTCTG

3. Differences in the expression of CsAOS in different tea plant varieties with suitability

The three leaves of different tea plant varieties with different suitability were picked, and the expression of allene oxide synthase gene CsAOS in each sample was detected by high-throughput sequencing.

Test results and analysis:

In Figure 2, it can be seen that the expression of the allene oxide synthase gene CsAOS in different tissues of Shuchazao is different, and its expression level is higher in two leaves and flowers, indicating that it plays a corresponding function in these tissues.

In Figure 3, we can see the difference in the expression of the allene oxide synthase gene CsAOS during black tea processing. Both transcriptome data and quantitative PCR data indicate that the expression of this gene gradually increases from withering to fermentation, indicating that The withering, rolling and fermentation of black tea promote the expression of this gene, and it also indicates that this gene is involved in the production process of black tea.

Fig. 4 is the expression of CsAOS in different suitable tea plant varieties according to the embodiment of the present invention. The results showed that the expression level of CsAOS in most of the tea tree varieties suitable for making black tea and dark tea was significantly higher than that in the tea tree varieties suitable for making green tea. Therefore, the expression of CsAOS gene can be used as a parameter to judge the suitability of tea varieties for making green tea and black tea.

Example 2 In vitro oligonucleotide antisense inhibition experiment

1. Design and synthesize oligonucleotide antisense primers according to the AOS predicted sequence. The design is completed on the website http://sfold.wadsworth.org/cgi-bin/soligo.pl . The primer sequences are as follows:

asODN001041-1 CTGTTGAGTGGTATTTTTCG

asODN001041-2 GTTGGGAGTGGAGTTTGGCT

asODN001041-3 GGGGTCCAGATAGGAGAGGA

asODN001041-4 GTATTGTGATGGCACGGGCG

2. Dissolve with 80mM sucrose solution to prepare inhibition buffer, blank is sucrose solution;

3. Use scissors to cut one bud and two leaves that are basically the same size, bright in color, healthy in color, and free of insects and diseases, and insert them into a 96-well plate with buffer. Make sure that the tail of one bud and two leaves is submerged in the buffer.

4. Put the 96-well plate into a lighted incubator for 16h of light/8h of darkness, and the temperature of the incubator is 28°C.

5. Take the treated 3d primer samples and blank samples after treatment.

Test results and analysis:

In Figure 5, it can be seen that the expression level of the allene oxide synthase gene CsAOS in the in vitro oligonucleotide antisense inhibition experiment was significantly inhibited compared with the control, and the inhibition rate was close to 2/3.

In Figure 6, we can see the measurement results of jasmonic acids after 3d antisense inhibition by oligonucleotides in vitro. It can be seen that inhibiting the expression of CsAOS can significantly reduce the jasmonic acid presupposition OPDA and the jasmonic acids JA, JA-Ile etc. content. Compared with the control, more than 50% of JA was reduced; at the same time, the content of salicylic acid SA was reduced.

In Figure 7, it can be seen that the in vitro oligonucleotide antisense inhibition experiment was performed after 3 days of volatiles determination, and the results showed that heptaldehyde, hexanal, 2-hexenal, 2,4-hexadiene were inhibited in vitro after CsAOS. Aldehyde, 2,4-heptadienal, 2-hexen-1-ol acetate, 2-ethylhexanol, benzyl alcohol, linalool, phenethyl alcohol, nonenal, nonanol, geraniol , 3-hexenyl ester hexanoic acid, 3-hexenoic acid-3-hexenyl ester, hexanoic acid-hexyl ester content increased, especially the content of hexanal, 2-hexenal, compared with the control increased by about 18 times and 8 times, respectively.

Example 3 Cloning of allene oxide synthase CsAOS gene and transient expression in vitro

1. Design specific primers, the underlined part is the restriction enzyme cleavage site, and the primer sequences are shown in SEQ ID NO.3 and SEQ ID NO.4:

SEQ ID NO. 3: TGGATCC ATGGCTTTTACTTCTCTAGC BamH1

SEQ ID NO. 4: C AAGCTT TCAAAAACTAGCTCTCTTGACG Hind3

2. According to the instructions of the plant total RNA extraction kit and the first-strand cDNA synthesis kit, the total RNA of the tea tree sample was extracted, and reverse transcribed into cDNA.

3. Using the reverse transcribed cDNA as a template, use the above-mentioned specific primers for amplification, PCR system: 2 μl forward primer, 2 μl reverse primer, 2 μl template, 25 μl MIX, ddH ₂ O19 μl liters; PCR program: the first step: 94°C for 5min. The amplification procedure was pre-denaturation at 98°C for 10s, denaturation at 98°C for 10s, annealing temperature at 57°C for 30s, extension at 68°C for 1 min, 35 cycles, and further extension at 68°C for 5 min. The obtained PCR products were stored at 4°C.

4. The PCR product was purified by a PCR purification kit, and then connected to pGEM-T Easy, and then transformed into DH5α. The colony PCR verification, gel running verification and sequencing verification were carried out as follows. The positive colonies were obtained, the T vector (CsAOS-pGEM) containing the CsAOS gene was obtained, and the DH5α Escherichia coli containing the CsAOS sequence of the allene oxide synthase gene was obtained.

Ligation conditions: ligation overnight at 4°C.

Conversion system:

The obtained ligation product was added to 50 μL of DH5α, gently mixed and placed in ice for 30 min; heat-shocked in a 42°C water bath for 45 sec, immediately placed in ice, and ice-bathed for 3 min; 1 mL of fresh LB medium was added, 37 ℃, 200 rpm shaking culture for 1 hour; centrifuge the cultured bacterial liquid, retain 300 μL of supernatant, resuspend and mix, plate, and culture at 37 ℃ overnight.

5. Design specific primers whose primer sequences are as follows:

F:GGGGACAAGTTTGTACAAAAAAGCAGGCTTCATGGCTTTTACTTCTCTAGC

R:GGGGACCACTTTGTACAAGAAAGCTGGGTTCAAAAACTAGCTCTCTTGACG

6. Using the CsAOS-pGEM obtained in step 4 as a template, using the primers in step 5 to amplify, using the method of homologous recombination (Gateway technology), through PCR verification, gel running verification and sequencing verification in turn, to construct CsAOS-PB2GW7. The fusion vector has a strong 35S promoter, and CsAOS-PB2GW7 was transformed into Agrobacterium GV3101 by freezing transformation.

Transient expression of target genes

1. The recombinant bacteria CsAOS-PB2GW7 GV3101 and the control GV3101 obtained in the examples were selected and inoculated into 5ml LB liquid medium (100ug/ml gentamicin, 50ug/ml spectinomycin), and cultured with shaking at 28°C.

2. Transfer 1 ml of overnight cultured Agrobacterium to 25 ml of LB liquid medium (with the same antibiotics as in 1, plus autoclaved acetosyringone).

3. Detect the OD600 value of the bacterial liquid cultured overnight.

4. Collect bacteria at 5000 g for 15 minutes, and resuspend the bacteria with resuspension solution (10 mM MgCl2, 10 mM 2-(N-morpholino)ethanesulfonic acid (pH 5.6), 100 uM acetosyringone), and the final OD600 is 0.4.

5. Inject tobacco after 2-3 hours at room temperature.

6. Put the infection solution into a 5ml syringe, press the reverse plate of the syringe with your thumb to inject the solution from the lower epidermis of the leaf into the tobacco leaf (do not use the cotyledon). After injection, the tobacco leaves will appear moist.

7. Three days after injection, the injected leaves were removed for analysis of volatiles and jasmonic acids.

Test results and analysis:

In FIG. 8 , the expression level of the in vitro transiently expressed allene oxide synthase gene CsAOS can be seen, and the results show that the expression level of CsAOS in the transiently expressed tobacco is increased by 9 times.

FIG. 9 is the measurement result of jasmonic acids after transient expression in vitro for 3 days according to the embodiment of the present invention. The results showed that transient expression significantly increased the content of jasmonic acids, such as JA and MeJA increased by about 20% and 29%, respectively.

Detection method involved in the present invention

In Vitro Oligonucleotide Antisense Inhibition Samples for Volatiles Assays:

1. The determination of volatiles was carried out by headspace solid-phase microextraction (HS-SPME) combined with gas mass spectrometry (GC-MS) (Agilent 7890A). The extraction head of PDMS (Supelco) extracted the volatiles of the antisense oligonucleotide antisense samples and blank samples in vitro, extracted at 60 °C for 1 h, then took out the extraction head and inserted it into the GC-MS injection port for desorption for 5 minutes, and started the instrument to collect data at the same time. .

2. GC-MS detection conditions

Chromatographic conditions: the inlet temperature is 240 °C, the carrier gas is high-purity nitrogen, the purity is greater than 99.99%, the flow rate is 0.8ml/min, and the sample is splitless; the chromatographic column model is HP-5MS quartz capillary column (60m×0.32mm× 0.25um); the column temperature starts at 40°C, holds for 3 minutes, rises to 90°C at 2°C/min, holds for 5 minutes, then rises to 160°C at 3°C/min, and finally rises to 250°C at 10°C/min, Hold for 5min.

Mass spectrometry conditions: ion source was EI source, ion source temperature was 230 °C, electron energy was 70 eV, quadrupole temperature was 150 °C, interface temperature was 280 °C, electron multiplier voltage was 1680 V, and the scanning range m/z was 35-350 amu.

Determination of jasmonic acids:

The samples were ground into powder in liquid nitrogen and extracted in the dark as follows.

The extraction buffer _formula was prepared, methanol:ddH2O:acetic acid=80:19:1 (V:V:V). Weigh about 0.1 g of the sample into a 2 ml centrifuge tube, add 750 μl of extract, invert and mix, and place on ice. This process needs to be carried out in the dark. Then, at 4°C, rotary extraction was performed for more than 16 h in the dark. Centrifuge at 13,000 rpm for 10 min at 4°C, suck the supernatant into a new centrifuge tube, add 750 μl of extract again, extract at 4°C in the dark for more than 16h, centrifuge, and combine the two supernatants. Filter through a 0.22um membrane (organic membrane), slowly dry with nitrogen, add 200 μl methanol, invert several times and dissolve at 4°C for 3 to 6 hours. (This process is carried out in the dark). After centrifuging the lysate at 4°C and 13000rpm for 15min, gently pipette 180μl of the supernatant into the inner tube, and place it in a special sample bottle for mass spectrometry to wait for on-board detection.

The hormone samples were analyzed by LC-MS, the liquid chromatograph was Shimadzu LC-20AD type, and the quantitative analysis was performed by the external standard method.

The detection method is as follows:

The liquid chromatography adopts a binary solvent system. The mobile phase is methanol in solution A, and 0.05% formic acid aqueous solution in solution B. Eclipse plus C18 (5μm, 2.1*150mm) chromatographic column is selected, the control flow rate is 300uL/min, and the column temperature is 30 °C, 10 μL per injection. Gradient elution was used, starting with a gradient of 10% methanol for 2 min, gradually increasing to 90% for 5 min at 10 min. At 15.1 min, methanol was reduced to the initial gradient and held for 7 min.

Table 1 Liquid-phase elution conditions of plant hormones

The mass spectrometry conditions are as follows: the ion source is an electrospray ion source (ESI), the voltage of the ion source is -4.5KV, the temperature of the ion source is 500°C, the auxiliary heating gas gas2 (50psi) is N2, and the atomizing gas gas1 (60psi) is used. and curtain gas (30psi), the test object is in negative ion mode. Standard mixtures were separated by reversed-phase liquid chromatography and quantified by tandem triple quadrupole mass spectrometry in multiple reaction detection (MRM) mode with a scan time of 50 ms.

The above descriptions are only preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements and improvements made within the spirit and principles of the present invention shall be included in the protection of the present invention. within the range.

sequence listing

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<211> 29

<212> DNA

<213> Artificial sequences (Hind3)

<400> 4

caagctttca aaaactagct ctcttgacg 29

Claims (10)

1. An allene oxide synthetase gene CsAOS, the nucleotide sequence of which is shown in SEQ ID NO.1, or a DNA sequence which is hybridized with the DNA sequence limited by SEQ ID NO.1 and encodes the protein with the same function; or DNA molecule which has more than 90% of homology with the DNA sequence limited by SEQ ID NO.1 and codes the same functional protein.

2. An allene oxide synthetase has an amino acid sequence shown in SEQ ID NO.2, or an amino acid sequence which is obtained by substituting and/or deleting and/or adding a plurality of amino acid residues in the sequence shown in SEQ ID NO.2 and has the same protein function; or

Derived from the amino acid sequence shown in SEQ ID NO.2, has more than 98 percent of homology and has the same protein function.

3. Use of the allene oxide synthase gene CsAOS according to claim 1 for the preparation and/or use as an allene oxide synthase.

4. Use of the allene oxide synthase gene CsAOS according to claim 1 for the regulation of volatile substance content in tea.

5. Use according to claim 4, characterized in that: the volatile substances in the tea leaves are at least one of heptanal, hexanal, 2-hexenal, 2, 4-hexadienal, 2, 4-heptadienal, 2-hexen-1-ol acetate, 2-ethylhexanol, benzyl alcohol, linalool, phenethyl alcohol, nonenal, nonanol, geraniol, 3-hexenyl ester hexanoic acid, 3-hexenyl ester or hexanoic acid-hexyl ester.

6. The use of the allene oxide synthase gene CsAOS according to claim 1 for the regulation of jasmonates content and/or salicylic acid content in tea.

7. Use according to claim 6, characterized in that: the jasmonic acid substance is at least one of jasmonic acid, jasmonic acid amino acid and methyl jasmonate.

8. The use of the allene oxide synthase gene CsAOS according to claim 1 for adjusting the content of jasmonic acid precursor OPDA in tea.

9. Use of the allene oxide synthase gene CsAOS according to claim 1 as a marker for screening tea varieties suitable for making green tea and/or tea varieties suitable for making black tea or black tea.

10. A method for promoting the expression level of allene oxide synthase gene CsAOS in tea leaves is characterized in that: comprises at least one treatment mode of withering, rolling and fermenting treatment steps of the tea leaves.

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