CN109182350B - Application of Maize Zm675 Gene in Plant Quality Improvement - Google Patents

Abstract

The invention relates to application of a corn Zm675 gene in plant quality improvement. The invention discloses an application of a Zm675 gene, a Zm675 protein or an expression cassette or a vector containing the Zm675 gene in improving the content of lysine and/or protein in plants. Zm675 gene was cloned in maize, and transgenic maize into which Zm675 gene was introduced was constructed. The lysine content and protein content in the seeds of the transgenic corn for enhancing Zm675 gene expression are obviously improved, and the number of protein bodies in corn endosperm cells is obviously increased. Meanwhile, the excellent properties of high protein and high lysine of the transgenic corn introduced with the Zm675 gene can be stably inherited. The Zm675 gene has important application value for improving the content of lysine and protein in plant grains and improving the nutritional quality of plants.

Description

Application of Maize Zm675 Gene in Plant Quality Improvement

technical field

The invention relates to the fields of biotechnology and genetics and breeding, in particular to the application of maize Zm675 gene in plant quality improvement.

Background technique

Corn is an important food crop and feed crop, but the content of protein and certain essential amino acids in corn kernels is low, which limits the nutritional value of corn to a certain extent. Lysine is the first limiting amino acid in corn grains. Therefore, increasing the content of protein and essential amino acids in grains, especially lysine, can improve the nutritional quality of corn. In 1964, the discovery of the opaque2 maize mutant opened a precedent for the improvement of the nutritional quality of maize. The O2 gene encodes a bZIP-like transcription factor that can regulate the synthesis of gliadin, and the mutation of the O2 gene leads to the lack of lysine in maize endosperm. The proportion of gliadin decreased, and the proportion of lysine in endosperm increased (Schmidt et al., 1990). The lysine content of the maize endosperm of opaque-2(o2) mutants is 70%–100% higher than that of common maize (Mertz et al., 1964), but due to the opaque soft endosperm of the kernel, its yield is low, There are many defects such as low seed germination rate, poor seedling growth, susceptibility to pests and diseases, easy mildew, intolerance to storage and poor processing quality. QPM (high-quality protein maize) cultivated by conventional breeding using endosperm modified genes overcomes the shortcomings of opaque2 mutants. QPM not only has high lysine content, but also has a hard endosperm and excellent agronomic traits (Gibbon and Larkins, 2005). It has obvious nutritional advantages and economic value. But QPM corn still has some flaws and deficiencies: (1) Compared with common corn, the yield of QPM corn is lower; (2) QPM production faces severe biotic (disease and insect pests) and abiotic (drought, heat, low soil pH, low soil nitrogen, etc.) constraints. The characteristics of long breeding cycle, low efficiency and poor predictability increase the difficulty of breeding high-resistant QPM varieties. (3) Since the o2 allele that provides QPM traits is recessive, when QPM maize is inoculated with normal endosperm maize pollen, the excellent genetic traits of QPM will be lost. Therefore, QPM production requires isolation, which increases the demands on the growing environment (Lilianee et al., 2017).

The development of genetic engineering technology provides a new idea for obtaining corn with high lysine content. On the one hand, the content of free lysine in corn grains can be increased by regulating the lysine synthesis and metabolism pathway. Reyes et al. used RNA interference technology to inhibit the activity of LKR/SDH, a key enzyme in the process of lysine catabolism, and increased the content of free lysine in corn grains (Reyes et al. 2009). However, the accumulation of excess free lysine will affect the development of corn kernels, and the instability of free lysine makes it easily damaged during processing (Wenefrida et al., 2013).

On the other hand, by inhibiting the expression of 19-kD or 22-kD α-gliadin and reducing the content of 19kD or 22kD gliadin in the grain, corn with higher lysine content can be obtained (Segal et al. 2003; Huang et al. 2004; Huang et al. 2005; ) However, this maize has the same grain phenotype as the o2 mutant, producing a floury endosperm, limiting its application in production (Wu and Messing, 2012).

The expression of high-quality lysine-rich protein gene in seeds is a new way to obtain high-lysine-content maize and overcome the soft endosperm problem. The key to this method is to obtain high-quality high-lysine content protein. genetic resources. At present, a class of lysine-rich microtubule-binding protein genes from dicotyledonous plants are mainly used in maize quality improvement research (Chang et al., 2015; Lang et al., 2004; Liu et al., 2015 ; Yu et al., 2004; Yue J, et al., 2014).

SUMMARY OF THE INVENTION

In order to solve the problems existing in the prior art, the purpose of the present invention is to provide the application of the maize Zm675 gene in plant quality improvement.

In the process of researching plant quality improvement, the inventor found that the Zm675 gene of maize has the function of increasing the lysine content and protein content of plants. The function of the maize Zm675 gene is currently unknown, and the lysine content of the encoded protein is 18.56%. However, in addition to the relatively high lysine content of the encoded protein of the Zm675 gene, the inventors also found that increasing the expression of the Zm675 gene can not only significantly increase the lysine content in plants, but also significantly increase the protein content in plants. At the same time, the synthesis and accumulation of protein bodies in plant seeds also increased significantly. Therefore, it is speculated that the Zm675 gene is involved in regulating the formation of protein bodies and affects the accumulation of other proteins including proteins with high lysine content in seeds, thereby increasing the lysine and protein content of seeds and improving plant seeds. quality.

First, the present invention provides the application of Zm675 gene, Zm675 protein or expression cassette or vector containing Zm675 gene in increasing plant lysine and/or protein content.

Or, the application of Zm675 gene, Zm675 protein or expression cassette or vector containing Zm675 gene in improving plant quality.

Or, the application of Zm675 gene, Zm675 protein or expression cassette or vector containing Zm675 gene in the preparation of transgenic plants with improved quality.

The Zm675 gene of the present invention has the following nucleotide sequence:

(1) the nucleotide sequence as shown in SEQ ID NO.2;

(2) a nucleotide sequence encoding a protein with the same function obtained by the substitution, deletion or insertion of one or more bases in the nucleotide sequence shown in SEQ ID NO.2;

(3) A nucleotide sequence capable of hybridizing with the nucleotide sequence described in (1) or (2) and encoding a protein with the same function under stringent conditions.

The Zm675 protein of the present invention has the following amino acid sequence:

(1) amino acid sequence as shown in SEQ ID NO.1;

(2) The amino acid sequence of a protein with the same function obtained by substitution, deletion or insertion of one or more amino acids in the amino acid sequence shown in SEQ ID NO.1.

Preferably, the application of the present invention is to increase the expression of the Zm675 gene;

More preferably, the increase in the expression level of the Zm675 gene is to introduce the Zm675 gene into a plant.

The Zm675 gene provided by the present invention is derived from maize. Therefore, the increase in the expression level of the Zm675 gene can be achieved by increasing the expression level of the Zm675 gene in maize or by heterologously expressing the Zm675 gene in other plants by transgenic methods.

The plants of the present invention can be either monocotyledonous or dicotyledonous plants.

Preferably, the plant of the present invention is corn.

Further, the present invention provides a method for preparing a transgenic plant with increased lysine and/or protein content, the method is to increase the expression level of the Zm675 gene.

Preferably, the improving the expression level of the Zm675 gene is to introduce the Zm675 gene into a plant; the Zm675 gene has the following nucleotide sequence:

(1) the nucleotide sequence as shown in SEQ ID NO.2;

(2) a nucleotide sequence encoding a protein with the same function obtained by the substitution, deletion or insertion of one or more bases in the nucleotide sequence shown in SEQ ID NO.2;

(3) A nucleotide sequence capable of hybridizing with the nucleotide sequence described in (1) or (2) and encoding a protein with the same function under stringent conditions.

Stringent conditions were hybridization in 0.1 % SDS in 0.1 x SSPE or 0.1 % SDS in 0.1 x SSC at 65°C and membrane washing with this solution.

Specifically, the method for preparing a transgenic plant with increased lysine and/or protein content comprises the following steps:

(1) construct an expression vector containing the Zm675 gene;

(2) Introducing the expression vector into a plant to obtain a transgenic plant.

As a preferred embodiment of the present invention, the expression vector containing the Zm675 gene is constructed based on the plant expression vector pSB130-F128 (ZL201310398102.9);

The expression vector includes two T-DNAs: wherein, T-DNA1 includes the Zm675 expression cassette driven by the millet seed specific promoter F128 (CN101063139A); T-DNA2 includes HPT (hygromycin phosphate) driven by the CaMV35S promoter. transferase gene) expression cassette.

The introduction of the expression vector into a plant is a method by Agrobacterium infection.

The beneficial effects of the present invention are as follows: the present invention clones the Zm675 gene in maize, and constructs the transgenic maize by increasing the expression level of the Zm675 gene. The content of lysine in the seeds of transgenic maize with enhanced Zm675 gene expression was increased by more than 20%, up to 41.17%, and the content of protein was up to 12.84%. The number of starch grains in maize endosperm cells was significantly reduced, and the number of protein bodies was significantly increased. At the same time, the excellent traits of high protein and high lysine in the transgenic maize introduced into Zm675 gene can be stably inherited; moreover, the introduction of Zm675 gene does not affect the endosperm traits and germination rate of maize seeds, and the endosperm phenotype of transgenic maize seeds is normal. , no powdery endosperm appeared, and the germination rate of seeds was not significantly different from that of the wild type. Therefore, Zm675 gene can be used to increase the content of lysine and protein in plant grains and improve the nutritional quality of plants.

Description of drawings

FIG. 1 is a diagram showing the construction strategy of the plant expression vector containing the Zm675 gene in Example 2. FIG.

FIG. 2 is the PCR detection of HPT in some corn regenerated plants of TO generation in Example 4. FIG. Among them, 3-5 are transgenic plants F4, F10 and F11, respectively.

FIG. 3 shows the PCR detection of Zm675 gene in some maize plants of T1 generation in Example 4. FIG. Among them, 3-5 were transgenic lines F4, F10 and F11, respectively.

Figure 4 is the expression level analysis of the Zm675 gene in the immature seeds of R1 generation maize in Example 6; Actin is used as the internal reference gene, the data is the average value of three biological replicates, and the significance analysis is based on the t test (*p<0.05, * *p<0.01, ***p<0.001).

FIG. 5 is a diagram showing the results of transmission electron microscopy of immature maize endosperm in Example 7, wherein Bar=5 μm; PB represents protein body; S represents starch grain.

Figure 6 shows the grain phenotype and germination rate of the transgenic maize in Example 8, wherein A is the grain phenotype, and the 1st, 3rd, 4th, and 6th rows from top to bottom are the grains photographed under white light, and the 2nd and 5th rows For the corn kernels photographed under transmitted light, the 1st, 2nd, 4th, and 5th rows are complete seeds, and the 3rd and 6th rows are the cross-sections of the same parts of the seeds; B is the seed germination rate, and the seed germination experiment was repeated three times. The data are Average of three experiments, error bars are standard deviation.

Detailed ways

The preferred embodiments of the present invention will be described in detail below with reference to the examples. It should be understood that the following examples are given for illustrative purposes only, and are not intended to limit the scope of the present invention. Those skilled in the art can make various modifications and substitutions to the present invention without departing from the spirit and spirit of the present invention.

The experimental methods used in the following examples are conventional methods unless otherwise specified.

The materials, reagents, etc. used in the following examples can be obtained from commercial sources unless otherwise specified.

Example 1 Cloning and sequence analysis of Zm675 gene

The maize inbred line B73 was used as material to extract total RNA from leaves, and cDNA was synthesized by reverse transcription with Oligo dT as primer.

Based on the maize genome sequence (maizeGDB: GRMZM2G051675), primers 675-F1 (SEQ ID NO. 3): 5'-CACCAGAGGCGATGTCGTC-3' and 675-R1 (SEQ ID NO. 4): 5'-TGGTGAGAGCATCTTCAGTC-3' were designed to cDNA As a template, PCR amplification was performed. Reaction conditions: 95°C, 30sec; 58°C, 30sec, 72°C, 30sec, 30 cycles. The amplified product was ligated with T vector pMD-19T and transformed into E. coli DH5α to obtain a recombinant plasmid. The sequencing results showed that the full-length coding sequence of cloned Zm675 was 669bp (as shown in SEQ ID NO.2), encoding 222 amino acid residues (as shown in SEQ ID NO.1), and the lysine content of the encoded protein was was 18.56% (w/w).

Example 2 Construction of expression vector pSB130-675

Zm675 was excised from the above-mentioned T vector with BamHI and HincII, and the plant expression vector pSB130-F128 (ZL201310398102.9) was double digested with BamHI and Sma I, and the Zm675 fragment was connected to the vector to construct a recombinant plasmid pSB130-675 (Fig. 1). The plasmid includes two T-DNAs. T-DNA1 includes the Zm675 expression cassette driven by the foxtail millet seed specific promoter F128 (CN101063139A); T-DNA2 includes the HPT (hygromycin phosphotransferase gene) expression cassette driven by the CaMV35S promoter. After the pSB130-675 vector was successfully constructed, it was transferred into Agrobacterium LBA4404 for genetic transformation of maize. Agrobacterium transformation is performed using routine methods in the art.

Example 3 Genetic transformation of young maize embryos

Preparation of Agrobacterium liquid: Agrobacterium LBA4404 (pSB130-675) was cultured on YEB solid medium (containing Sm 125 μg/mL, Kan 100 μg/mL) at 28°C for two days and then the cells were collected. Suspend the bacteria with the infection medium to an OD ₆₀₀ of 0.6-0.8, shake the bacteria at 28°C and 75 rpm in the dark for 2-4 hours, and dilute the shaken bacterial solution with the infection medium to an OD ₆₀₀ of 0.3-0.4 for later use . Taking the maize inbred line 178 as the transformation receptor, the young embryos (about 1.5-2.0 mm in size) 12 days after pollination of the 178 inbred line were stripped, soaked and rinsed twice with the infection medium without Agrobacterium , pour off the remaining medium, infect the immature embryos with the prepared Agrobacterium tumefaciens for 15 minutes, then place the infected immature embryos on sterile filter paper to dry, transfer to the co-cultivation medium, and place the scutellum toward the Incubate at 20°C for 3 days in the dark. After 3 days, the immature embryos were transferred to recovery medium, and after 7 days of dark culture at 28°C, they were transferred to selection medium containing hygromycin, cultured at 28°C in the dark, and subcultured once every two weeks (the selection condition was 5 mg/mL of hygromycin). L, 15mg/L, 20mg/L, 20mg/L of hygromycin). The screened callus was transformed into recovery medium and cultured in dark conditions for 7-10 days. Then the callus was transferred to the differentiation medium, cultured in light, and the photoperiod was light/dark for 16h/8h, and subculture every two weeks. When the differentiated seedlings grow to 3-4cm, the seedlings are cut from the resistant callus, transferred to the rooting medium to induce rooting, transferred to the greenhouse when the root system of the seedlings is relatively developed, and planted in small pots after hardening. , and then transferred to large pots in the greenhouse for further cultivation, and the seeds were harvested after maturity. See Table 1 for the medium formula of each stage of maize transformation.

Table 1 Maize transformation medium formula (1L) at each stage

Example 4 PCR detection of genomic DNA of transgenic maize

PCR detection: Extract the genomic DNA of the regenerated plant obtained in Example 3, and use primers HPT3 (SEQ ID NO.5): 5'-TCGGCTCCAACAATGTCCTG-3' and HPT4 (SEQ ID NO.6): 5'-CGGTCGGCATCTACTCTATTCC-3' to carry out PCR amplification, detection of the screening marker gene HPT. Positive plants can amplify the target band of about 450bp, and the PCR detection results of some materials are shown in Figure 2.

The T0 generation maize with positive PCR detection was planted, and the T1 generation transgenic maize was obtained by self-pollination. Due to the independent integration of the two T-DNAs on the vector, there will be separation in the offspring, so it is detected whether the Zm675 gene is transferred in the offspring. There is an endogenous Zm675 gene in maize, but the endogenous Zm675 gene contains seven introns, and the pSB130-675 expression vector only includes the CDS of Zm675, so the cross-intron primer 675-196-F (SEQ ID NO.7): 5'-ACAAGGCCCACAAGGACG-3' and 675-651-R (SEQ ID NO.8): 5'-CAGGGTCACCACTCAGAAATGA-3', the target gene was detected by PCR. Reaction conditions: 95°C, 30sec; 56°C, 30sec, 72°C, 30sec, 30 cycles. Primer 675-196-F/675-651-R spanned the second to seventh intron, amplified 456 bp on CDS, and amplified 2286 bp from genomic DNA as template; The time was controlled at 30sec, and the endogenous sequences on the genome could not be amplified due to the short extension time. The results showed that the specific sequence of the introduced Zm675 gene could be detected in the transgenic lines, but this band was not amplified in the wild-type maize genomic DNA (Figure 3).

Example 5 Determination of lysine and protein content in T1 generation corn seeds

The lysine content of the seeds of T1 generation transgenic maize was determined by the ninhydrin method (Yue J, et al., 2014). For each transgenic line, 15 plants were planted, and 5 plants with positive target gene PCR detection were selected for detection. The results showed that compared with the wild-type maize 178 inbred line planted at the same time, the lysine content in the seeds of the obtained transgenic lines was significantly increased to different degrees, and the lysine content of the F4, F10 and F11 lines was significantly increased. The content improvement rates were 29.41%, 20.58% and 41.17%, respectively. The protein content of corn seeds was determined by semi-micro Kjeldahl method (GB2905-82), and the conversion coefficient of nitrogen and protein content was 6.25. Compared with the wild type, the protein content of the seeds of F4, F10 and F11 transgenic lines were significantly increased in different degrees, and the increase rates were 9.65%, 5.50% and 12.84%, respectively (Table 2).

Table 2 Lysine and protein content of T1 generation transgenic maize seeds

Values are mean ± SD, and the significance analysis between each transgenic line and WT was performed according to t test (*p<0.05, **p<0.01, ***p<0.001)

The three lines F4, F10 and F11 were screened and monitored continuously for 5 generations, and 5 plants were selected from each transgenic line. The results showed that the lysine and protein contents of the seeds of the transgenic progeny lines ranged from T ₂ to T The _5th generation has different degrees of improvement, and the stability is better. In the T5 generation seeds, the lysine content of F4 and F10 increased by more than 20%, and the protein content increased by about 10%, while the lysine content of F11 increased by more than 40% and the protein content increased by more than 15% (Table 3). This indicated that the Zm675 gene was stable in inbred lines, and the excellent traits of high protein and high lysine could be inherited stably.

Table 3 The increase rate of lysine and protein content of transgenic maize progeny seeds

Example 6 Expression level analysis of Zm675 gene in T1 generation maize seeds

To verify the correlation between the increase of lysine and protein content in F4, F10 and F11 and Zm675, the transcription level of Zm675 in transgenic maize was detected. Select F4, F10 and F11 T1 generation PCR positive plants, select 6 plants for each line, extract the endosperm RNA 20 days after pollination, reverse transcribed into cDNA. Real-time PCR analysis was performed with cDNA as template and primers 675-196-F/675-651-R, and the average value of 6 plants was taken for each line. Compared with wild-type plants, the expression levels of Zm675 in transgenic lines F4, F10 and F11 were significantly increased (Fig. 4). Among them, the transcription level of Zm675 in the F11 line was the highest. Combined with the transgenic lines in Example 5 The analysis results of lysine content and protein content showed that the difference of lysine content and protein content of each transgenic line was caused by the difference of Zm675 gene expression. The protein content increased with the increase of Zm675 gene expression.

Example 7 Transmission electron microscope observation of immature endosperm of transgenic maize

The grains of T5 generation transgenic maize were selected 20 days after pollination, and about 2mm thick longitudinal slices were cut from the endosperm tissue. Then the slices were invaded into the fixative prepared in advance, and the fixed material was ultrathinly sliced, and the sliced slices were observed under a transmission electron microscope. In the endosperm of the wild-type 178 inbred line, the starch granules were larger in size, with white scattered texture, distributed more and densely; the protein bodies were distributed around the starch granules, in the shape of small gray dots. Compared with the wild type, the number of starch grains in maize endosperm cells of the three transgenic lines F4, F10 and F11 was significantly reduced, while the number of protein bodies was significantly increased and the distribution was dense (Fig. 5). The results showed that the Zm675 gene was involved in regulating protein bodies. Formation and accumulation affect the accumulation of various proteins in seeds, including proteins with high lysine content, thereby increasing the lysine and protein content of seeds and improving the quality of plant seeds.

Example 8 Analysis of Zm675 Transgenic Maize Grain Character and Germination Rate

The T ₂ generation transgenic maize seeds and wild-type maize seeds of F4, F10 and F11 were taken, and three seeds were randomly selected from each line. The results are shown in Figure 6A, compared with the wild type, there is no significant difference in the grain status of the transgenic lines; the endosperm phenotype of the transgenic maize is normal, and no powdery endosperm appears, indicating that the expression of the Zm675 gene does not affect maize grains. growth state. At the same time, germination experiments were carried out on maize seeds of three transgenic lines, F4, F10 and F11. Take wild type and F4, F10 and F11 three lines of T ₂ generation transgenic maize seeds, 100 seeds each, germinate for 5 days, and observe and count the germination rate. Data analysis showed that the seed germination rates of F4, F10 and F11 were similar to those of the wild type, all above 85% (as shown in Figure 6B), indicating that the expression of Zm675 did not affect the germination of maize seeds.

references

Chang, Y., Shen, E., Wen, L., Yu, J., Zhu, D., and Zhao, Q. (2015). Seed-SpecificExpression of the Arabidopsis AtMAP18 gene increases both lysine and totalprotein content in Maize .PLoS ONE 10, e142952.

Gibbon, B.C., and Larkins, B.A. (2005). Molecular genetic approaches to developing quality protein maize. Trends Genet. 21, 227-233.

Huang,S.,Adams,W.R.,Zhou,Q.,Malloy,K.P.,Voyles,D.A.,Anthony,J.,Kriz,A.L.and Luethy,M.H.(2004)Improving nutritional quality of maize proteins by expressing sense and antisense zein genes. J. Agric. Food Chem. 52, 1958–1964.

Huang, S., Kruger, D.E., Frizzi, A., D'Ordine, R.L., Florida, C.A., Adams, W.R., Brown, W.E., and Luethy, M.H. (2005). High-lysine corn produced by the combination of enhanced lysine biosynthesis and reduced zein accumulation. Plant Biotech J, 2005, 3:555–569

Lang, Z., Zhao, Q., Yu, J., Zhu, D., and Ao., G. (2004). Cloning of potato SBgLRgene and its intron splicing in transgenic maize. Plant Sci. 166, 1227-1233 .

Liliane N.T., Charles S.M., Eddy L.M.N., NoéW., (2017) Breeding for Quality Protein Maize (QPM) Varieties: A Review Agronomy. 7, 80; doi: 10.3390/agronomy7040080

Liu, C., Li, S., Yue, J., Xiao, W., Zhao, Q., Zhu, D., Yu, J. (2015). Microtubule-associated protein SBgLR facilitates storage protein deposition and its expression leads to lysine content increase in transgenic maizeendosperm.Int.J.Mol.Sci.16, 29772-29786.

Mertz, E.T., Bates, L.S., and Nelson, O.E. (1964). Mutant gene that changes protein composition and increases lysine content of maize endosperm. Science 145, 279-280.

Reyes, A.R., Bonin C.P., Houmard N.M., Huang S. and Malvar T.M. (2009). Genetic manipulation of lysine catabolism in maize kernels. Plant Mol Biol, 69(1):81-89

Schmidt, R.J., Burr, F.A., Aukerman, M.J., and Burr, B. (1990). Maizeregulatory gene opaque-2 encodes a protein with a "leucine-zipper" motif that binds to zein DNA.Proc.Natl.Acad.Sci.U.S.A. 87,46-50.

Segal, G., Song, R., and Messing, J. (2003). A new opaque variant of maize by a single dominant RNA-interference-inducing transgene. Genetics 165, 387-397.

Wenefrida, I., Utomo, H.S., and Linscombe, S.D. (2013). Mutational breeding and genetic engineering in the development of high grain protein content. J. Agric. Food Chem. 61, 11702-11710.

Wu, Y., and Messing, J. (2012). RNA interference can rebalance the nitrogen sink of maize seeds without losing hard endosperm. PLoS ONE 7, e32850.

Yu, J., Peng, P., Zhang, X., Zhao, Q., Zhu, D., Sun, X., Liu, J., and Ao, G. (2005). Seed-specific expression of the lysine-rich protein gene sb401 significantly increases both lysine and total protein content in maize seeds. FoodNutr. Bull. 26, 427-431.

Yue J, Li C, Zhao Q, Zhu D, Yu J. Seed-Specific Expression of a Lysine-Rich Protein Gene, GhLRP, from Cotton Significantly Increases the LysineContent in Maize Seeds. International Journal of Molecular Sciences 2014;15(4) :5350–5365.

The above are only the preferred embodiments of the present invention. It should be pointed out that for those skilled in the art, without departing from the technical principles of the present invention, several improvements and modifications can be made. These improvements and modifications It should also be regarded as the protection scope of the present invention.

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Claims (6)

1. The application of Zm675 gene, Zm675 protein or an expression cassette or a vector containing the Zm675 gene in improving the content of lysine and/or protein in plants, wherein the amino acid sequence of the Zm675 protein is shown as SEQ ID NO. 1; the nucleotide sequence of the Zm675 gene is shown in SEQ ID NO. 2.
2. The application of Zm675 gene, Zm675 protein or an expression cassette or a vector containing the Zm675 gene in improving the quality of plants, wherein the amino acid sequence of the Zm675 protein is shown as SEQ ID NO. 1; the nucleotide sequence of the Zm675 gene is shown in SEQ ID NO. 2; the plant quality is lysine and/or protein content.
3. Use of Zm675 gene, Zm675 protein, or an expression cassette or vector comprising Zm675 gene, wherein the amino acid sequence of Zm675 protein is represented by SEQ ID No.1, for the preparation of transgenic plants with improved lysine and/or protein content; the nucleotide sequence of the Zm675 gene is shown in SEQ ID NO. 2.
4. 4. The use of any one of claims 1 to 3, wherein the use is for increasing the expression level of Zm675 gene.
5. 5. The use according to claim 4, wherein the increase in the expression level of Zm675 gene is carried out by introducing Zm675 gene into a plant.
6. 6. Use according to any one of claims 1 to 3, wherein the plant is a monocotyledonous or dicotyledonous plant.

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