Application of overexpression OsCHS1 gene in promoting rice growth, improving rice phosphorus absorption and improving rice low-phosphorus resistance

Abstract

The invention discloses application of an overexpression OsCHS1 gene in promoting rice growth, improving rice phosphorus absorption and improving rice low-phosphorus resistance. The OsCHS1 gene is transferred into a rice wild type (WT) plant, so that a rice over-expression OsCHS1 plant is obtained. Under the control and phosphorus element reduction conditions, compared with WT, the OsCHS1 gene expression quantity, plant height, overground part fresh weight, root system fresh weight, overground part dry weight, root system dry weight, total root length and overground part phosphorus concentration of a rice over-expression OsCHS1 plant are remarkably increased. Therefore, rice growth and rice phosphorus absorption can be improved and promoted through an OsCHS1 gene overexpression method, and a foundation is laid for efficient utilization of phosphorus and high yield of rice.

Description

Technical Field

[0001] The present disclosure relates to the technical field of plant genetic engineering, and more specifically, to the application of overexpressing the OsCHS1 gene in promoting rice growth, improving phosphorus uptake by rice, and enhancing the low-phosphorus tolerance of rice. Background Art

[0002] Rice is one of the most important cereals for humans. More than half of the world's population depends on rice as the staple food. Ensuring rice production is crucial for food security.

[0003] Phosphorus is one of the three major nutrient elements for plants and is an essential element for crop yield formation. During crop cultivation, phosphorus fertilizer must be applied to supplement it. The phosphorus that plants absorb and utilize mainly comes from the soil, but 95% of the phosphorus in the soil exists in forms that are difficult for plants to absorb, resulting in phosphorus deficiency in approximately 74% of the cultivated land in China. To pursue high yields, the input of phosphorus fertilizer often exceeds the actual demand of crops, leading to an increase in soil phosphorus accumulation. A considerable proportion of phosphorus combines with Fe 3+ , Ca 2+ , Al 3+ in the soil to form compounds and lose effectiveness, reducing the utilization efficiency of phosphorus. As a result, the utilization rate of applied phosphorus fertilizer in the current season is only 5% - 25%. Rice is the most important food crop in China, accounting for more than half of the total grain output in China. Rice production has a large demand for phosphorus fertilizer, but the utilization rate of phosphorus fertilizer is relatively low. Some phosphorus is lost through surface runoff, causing agricultural non-point source pollution. Therefore, reducing the input of phosphorus fertilizer and improving soil phosphorus availability are the keys to increasing rice yield and technological breakthroughs.

[0004] In recent years, more and more attention has been paid to improving the phosphorus utilization efficiency of rice by gene overexpression. However, there are relatively few reports that can clearly promote phosphorus uptake by rice and improve rice dry weight and other indicators. Summary of the Invention

[0005] In view of this, the purpose of the present invention is to provide the application of overexpressing the OsCHS1 gene in promoting rice growth, improving phosphorus uptake by rice, and enhancing the low-phosphorus tolerance of rice.

[0006] The technical solution adopted by the present invention is as follows: The first aspect of the present invention provides the application of overexpressing the OsCHS1 gene in promoting rice growth, and the nucleotide sequence of the OsCHS1 gene is as shown in SEQ ID NO.1.

[0007] Furthermore, the above promotion of rice growth is reflected in any one or more of the following aspects: 1) Increasing the plant height of rice; 2) Increasing the fresh weight of the above-ground part of rice; 3) Increasing the fresh weight of the rice roots; 4) Increase the above-ground dry weight of rice; 5) Increase the root dry weight of rice; 6) Increase the total root length of rice.

[0008] The second aspect of the present invention provides the application of overexpressing the OsCHS1 gene in improving phosphorus uptake in rice, and the nucleotide sequence of the OsCHS1 gene is shown in SEQ ID NO.1.

[0009] Furthermore, the improvement of phosphorus uptake in rice as described above is reflected in increasing the above-ground phosphorus content of rice under low-phosphorus conditions.

[0010] The third aspect of the present invention provides the application of overexpressing the OsCHS1 gene in improving the low-phosphorus tolerance of rice, and the nucleotide sequence of the OsCHS1 gene is shown in SEQ ID NO.1. Brief Description of the Drawings

[0011] Figure 1 : The map of the recombinant overexpression vector constructed in the examples of the present invention and the identification results; Figure 2 : The phenotype map of rice and the relative expression level of the OsCHS1 gene; Figure 3 : The comparison diagram of plant height after overexpression of the OsCHS1 gene; Figure 4 : The comparison diagram of above-ground fresh weight after overexpression of the OsCHS1 gene; Figure 5 : The comparison diagram of root fresh weight after overexpression of the OsCHS1 gene; Figure 6 : The comparison diagram of above-ground dry weight after overexpression of the OsCHS1 gene; Figure 7 : The comparison diagram of root dry weight after overexpression of the OsCHS1 gene; Figure 8 : The comparison diagram of total root length after overexpression of the OsCHS1 gene Figure 9 : The comparison diagram of above-ground phosphorus concentration after overexpression of the OsCHS1 gene; Figure 10 : The comparison diagram of root phosphorus concentration after overexpression of the OsCHS1 gene. Detailed Embodiments

[0012] The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0013] Primer sequence information: 39216(+): 5’-Gagagaacacgggggactttgcaacatggcggcggcggtgacgg-3’, 39216(-): 5’-agtctccgtcgtggtctttgtaatcggcggcggcgccggcggtg-3’.

[0014] The pBWA(V)HS-ccdB-3xflag-2 vector was provided by Wuhan Boyuan Biotechnology Co., Ltd.

[0015] Example 1: Using the cDNA of Nipponbare rice genome as a template, the OsCHS1 gene (SEQ ID NO.1) was amplified by PCR with primers 39216(+) and 39216(-); the reaction system for PCR amplification was 50 μL, including: 20 μL of Nuclease-free Water, 2 μL of each upstream and downstream primer, 25 μL of Biorun Pfu PCR Mix, and 1 μL of cDNA template; the reaction conditions for PCR amplification were: pre-denaturation at 94 °C for 5 min; denaturation at 94 °C for 30 s, annealing at 50 °C for 45 s, extension at 72 °C for 72 s, with 30 cycles; extension at 72 °C for 10 min. After purification of the PCR amplification product, it was ligated between the BsaI and Eco31I restriction sites of the pBWA(V)HS-ccdB-3xflag-2(2021) vector. The resulting recombinant vector was transformed into Escherichia coli E.coli, and positive clones were selected for detection. After correct sequencing, the vector was sent to Wuhan Boyuan Biotechnology Co., Ltd. and transformed into Agrobacterium, and then introduced into rice callus with Nipponbare rice as the background for genetic transformation. The transgenic rice obtained by genetic transformation was identified by hygromycin, and then by quantitative PCR. The primer sequences for identification were: OsCHS1F: 5’-ACTGCGTGTACCAGGCCGACTA-3’, OsCHS1R: 5’-TGGTGCAGAAGACGAGGTGGGT-3’. Finally, 2 homozygous OsCHS1 overexpression lines (OsCHS1 OE-1 and OsCHS1 OE-2) were obtained. Compared with wild-type rice, the relative expression level of the OsCHS1 gene in the OsCHS1 overexpression lines increased by about 1000 times ( Figure 2 ).

[0016] Example 2: After culturing and growing wild-type rice Nipponbare and OsCHS1 overexpression lines for 21 days under normal phosphorus (i.e., control, phosphorus concentration of 300 μM) and low phosphorus (phosphorus concentration of 2 μM) conditions respectively, samples were taken and plant height, above-ground fresh weight, root fresh weight, above-ground dry weight, root dry weight, total root length, above-ground phosphorus concentration, and root phosphorus concentration were measured respectively.

[0017] The results showed ( Figures 3 to 10 ), whether under control conditions or low phosphorus conditions, the plant height, above-ground fresh weight, root fresh weight, above-ground dry weight, root dry weight, and total root length of the OsCHS1 overexpression lines were significantly higher than those of wild-type rice Nipponbare; under low phosphorus conditions, the above-ground phosphorus concentration of the OsCHS1 overexpression lines was significantly higher than that of wild-type rice Nipponbare. Among them, under control conditions, compared with wild-type rice Nipponbare, the plant heights of OsCHS1 OE-1 and OsCHS1 OE-2 increased by 9.4% and 12.8% respectively, the above-ground fresh weights increased by 35.1% and 68.7% respectively, the root fresh weights increased by 25.2% and 53.5% respectively, the above-ground dry weights increased by 29.6% and 54.9% respectively, the root dry weights increased by 23.8% and 46.3% respectively, and the total root lengths increased by 40.1% and 48.5% respectively. Under low phosphorus conditions, compared with wild-type rice Nipponbare, the plant heights of OsCHS1 OE-1 and OsCHS1 OE-2 increased by 14.3% and 14.3% respectively, the above-ground fresh weights increased by 59.6% and 38.2% respectively, the root fresh weights increased by 87.7% and 55.3% respectively, the above-ground dry weights increased by 63.6% and 47.9% respectively, the root dry weights increased by 91.5% and 66.1% respectively, the total root lengths increased by 51.2% and 27% respectively, and the above-ground phosphorus concentrations increased by 35.3% and 35.3% respectively.

Claims

1. Overexpression OsCHS1 The application of the gene in promoting rice growth is characterized by: Said OsCHS1 The nucleotide sequence of the gene is shown in SEQ ID NO.

1.

2. The use according to claim 1, characterized in that: The promotion of rice growth is embodied in any one or more of the following aspects: 1) Increase rice plant height; 2) Increase the fresh weight of aboveground rice; 3) Increase the fresh weight of rice roots; 4) Increase the aboveground dry weight of rice; 5) Increase the dry weight of rice roots; 6) Increase the total root length of rice.

3. Overexpression OsCHS1 The application of the gene in improving phosphorus absorption in rice is characterized by: Said OsCHS1 The nucleotide sequence of the gene is shown in SEQ ID NO.

1.

4. The use according to claim 3, characterized in that: The improvement of phosphorus absorption of rice is reflected in increasing the phosphorus content of the aboveground part of rice under low-phosphorus conditions.

5. Overexpression OsCHS1 The application of the gene in improving the tolerance of rice to low phosphorus is characterized by: Said OsCHS1 The nucleotide sequence of the gene is shown in SEQ ID NO.1.

Images