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A kind of gene of Thalassiosira vissonii twpepc1 and its application and method for cultivating high-yielding rice

A technology of seaweed and seaweed, applied in the field of plant genetic engineering, can solve the problems that the yield of PEPC transgenic plants is not well improved, cannot exert due C4 effect, etc., and achieves overcoming the dependence of rosette structure , the effect of increasing activity and increasing yield

Active Publication Date: 2021-11-26
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] In previous reports, since exogenous PEPC genes are derived from two-cell C 4 In photosynthetic plants, such as corn, the effect of genes depends on its specific "rosette"-type organizational structure, which cannot exert the proper C in a single cell. 4 effect, and the gene may also have species regulation and modification specificity, so the yield of PEPC transgenic plants has not been well improved

Method used

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  • A kind of gene of Thalassiosira vissonii twpepc1 and its application and method for cultivating high-yielding rice
  • A kind of gene of Thalassiosira vissonii twpepc1 and its application and method for cultivating high-yielding rice
  • A kind of gene of Thalassiosira vissonii twpepc1 and its application and method for cultivating high-yielding rice

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Cloning of TwPEPC1 Gene from Thalassiosira vissonii

[0052] Material preparation: Thalassiosira vissonii, the culture conditions are temperature 22°C, light intensity 3000lux, light-dark ratio 16h / 8h.

[0053] Nutrient solution composition: NaNO 3 74.8mg / L, NaH 2 PO 4 4.4mg / L, vitamin mother solution 1ml (vitamin B1 0.1g / L, vitamin B12 0.5 mg / L, vitamin H 0.5mg / L, trace element mother solution 1ml (EDTA-Na 2 4.360 g / L, FeCl 3 .6H 2 O 3.150g / L, CuSO 4 .5H 2 O 0.010g / L, ZnSO 4 .7H 2 O 0.022g / L, CoCl 2 .6H 2 O 0.010g / L, MnCl 2 .4H 2 O 0.180g / L, Na 2 MoO 4 .2H 2 O 0.006g / L), pH=8.0, the culture medium was prepared with seawater, sterilized at 121°C for 15 minutes before use.

[0054] step:

[0055] (1) Extraction of Thalassiosira vissonii RNA:

[0056] Thalassiosira wilsonii RNA was obtained by using the method of Trizol Reagent (Invitrogen, Carisbad, CA, USA). The Thalassiosira wilsonii RNA was detected by 0.8% agarose gel electrophoresis and concentra...

Embodiment 2

[0064] Construction of Transformation Vector of Thalassiosira vissonii TwPEPC1 Gene

[0065] (1) Restriction vector pC1300s

[0066] The pC1300s vector was transformed on the basis of pCAMBIA1300 (the vector publicly used by CAMBIA Company), and the 35s promoter and 35s polyA vector map and the information of the multiple cloning site were respectively introduced at both ends of the multiple cloning site. figure 1 . pC1300s was digested with Kpn I, and dephosphorylation reaction was carried out after electrophoresis detection that the digestion was linear. The reaction system and conditions were referred to the instructions of Promega. Service Co., Ltd.) recovery, electrophoresis to detect the integrity of its digestion, and stored in a -20 ° C refrigerator.

[0067] (2) Enzyme digestion of TwPEPC1 cloning plasmid

[0068] The correct TA cloning plasmid was sequenced with Kpn I single enzyme digestion, and the digested product was separated by 0.8% agarose electrophoresis, ...

Embodiment 3

[0072] Agrobacterium-mediated genetic transformation

[0073] The Agrobacterium-mediated genetic transformation method of the present invention mainly refers to the method shown in the "Agrobacterium-mediated Genetic Transformation Operation Manual" issued by the State Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University (Lin Yongjun, etc., Agrobacterium-mediated Mudanjiang 8 No. Establishment of high-efficiency transgenic system, Acta Crops Sinica, 2002, 28(3): 294-300). The recipient of the transformation was the embryogenic callus induced from the mature seeds of the rice variety "Zhonghua 11". The hygromycin-resistant callus is obtained through precultivation, infection, cocultivation and screening, and then through differentiation, rooting, seedling training and transplanting to obtain transgenic plants.

[0074] (1) Abbreviation of reagents and solutions

[0075] The title and the abbreviation of the main reagent involved in the medium preparati...

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Abstract

The present invention provides a Thalassiosira wilsonii TwPEPC1 gene and a method for applying and cultivating high-yielding rice, which belong to the technical field of plant genetic engineering. The nucleotide sequence of the Thalassiosira wilsonii TwPEPC1 gene is shown in SEQ ID No.1 Show. The TwPEPC1 gene of Thalassiosira vissonii provided by the present invention has the effect of improving the yield of rice and overcomes the two-cell C 4 plant-derived C 4 Dependence of genes on rosette-shaped structures. The experimental results showed that the TwPEPC1 gene of Thalassiosira weiweieri was expressed in rice. Compared with the original rice plants, the PEPC enzyme activity of the transgenic plants was increased by 2 to 3 times, the photosynthesis rate was significantly improved, and the yield was significantly improved.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, and in particular relates to a Thalassiosira vissonii TwPEPC1 gene and a method for applying and cultivating high-yielding rice. Background technique [0002] With population growth, the total human demand for food continues to rise, while the area of ​​arable land decreases due to urbanization, which requires higher yields of crops (Surridge, 2002). C 4 plant than C 3 Plants have higher photosynthetic efficiency, greater biosynthesis, higher NUE and WUE and wider adaptability, but most important crops such as rice, wheat, potato and soybean are C 3 Plants (Von Caemmerer, 2003). Therefore, using transgenic technology in C 3 In-plant expression of C 4 Related genes (especially PEPC) also increase C 3 One of the potential strategies for crop yield (Zhang Qifa, 2009; Hibberd et al., 2008; Zhu et al., 2010a; Zhu et al., 2010b). [0003] Although most C 4 Plants are all roset...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/57C12N9/48C12N15/82A01H4/00A01H5/00A01H6/46
CPCA01H4/00C12N9/485C12N15/8261
Inventor 林拥军陈太钰
Owner HUAZHONG AGRI UNIV