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Application of 0scpy and its encoded protein in rice seeding

A technique for encoding protein and rice, applied to the field of application of an orphan gene Oscpy in rice and its encoded protein in rice seeding

Active Publication Date: 2022-05-17
NANCHANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In view of the complexity of genetic inheritance, the role of the rice orphan gene OsCPY, its encoded protein and its gene remains to be further developed.

Method used

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  • Application of 0scpy and its encoded protein in rice seeding
  • Application of 0scpy and its encoded protein in rice seeding
  • Application of 0scpy and its encoded protein in rice seeding

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Embodiment 1: Construction and genetic transformation of RNA interference carrier

[0069] (1) Obtaining the forward interference fragment: Extract the total RNA of the young panicle of Zhonghua No. 11, reverse it into cDNA, and use the designed primer pair 3 according to the Oscpy gene sequence: SEQ ID NO.3CPY-SIF22: 5'-CGCGGATCCTCCATCGTGGAGCCGGAGAA-3 ' (the underline is the restriction site of BamHI) and SEQ ID NO.4CPY-SIR22: 5'-GCCTAAGCTTCCCTGTAGAAACCTCCTGCTCCAG-3' (the underline is the restriction site of HindIII). The positive interference fragment (SEQID NO.5) of Oscpy was amplified by RT-PCR, and was used for the construction of the interference vector after sequencing verification.

[0070] (2) Construction of RNA interference vector: forward interference fragment (SEQ ID NO.5) and pYL vector (see figure 1 ) were subjected to double enzyme digestion, purification, and recovery with BamH I and Hind III enzymes, and then ligated with T4 DNase. The ligated product...

Embodiment 2

[0082] Example 2 Detection of Transgenic Positive Rice Plants and Obtaining of Oscpy Expression Down-regulated Plants

[0083] After the DNA of transgenic rice leaves was extracted by CTAB method, hygromycin phosphotransferase gene-specific primers (SEQ ID NO.9:5'-CTGAACTCACCGCGACGTCTGTC-3'; SEQ ID NO.10:5'-TAGCGCGTCTGCTGCTCCATACA-3' ) The rice that was amplified positive by PCR was determined to be the transgenic positive plant of the T0 generation; using the fluorescent quantitative PCR method, with Oscpy gene-specific primers SEQ ID NO.11:5'-AGGGCACTGATGCTAGAGGA-3'; SEQ ID NO.12:5 '-CTCATTGGCGCCGGATTTTC-3', using rice OsActin as an internal reference gene, its primer sequence is SEQ ID NO.13:5'-ACCTTCAACACCCCTGCTAT-3' and SEQ ID NO.14:5'-CACCATCACCAGTCC AAC-3' to detect the down-regulated expression of Oscpy , and obtain Oscpy gene down-regulated expression RNAi rice plants and T-DNA insertion mutant plants, such as image 3 As shown, compared with wild-type Zhonghua 11, w...

Embodiment 3

[0084] The observation of the growth of wild type and transgenic strain coleoptile under the condition of embodiment 3 flooding

[0085] Select mature and plump wild type Zhonghua 11 (ZH11), mutant (cpy), RNA interference (RNAi), 200 grains each, and first wet the rice seeds with sufficient distilled water to make the surface of the seeds hydrophilic. Put these wet seeds into a glass bottle marked with the experiment date and seed name, add an equal volume and sufficient amount of distilled water to germinate the seeds under completely submerged conditions, put them into day / night for 12 hours, and the temperature is 32°C during the day Germinate and cultivate in an incubator at 28°C in the dark, with a relative humidity of 60%, and a light intensity of 3000Lx. Replace the distilled water at the same temperature under the conditions of rice every 8 hours, and clean the seed metabolites in the glass bottle to avoid the poisoning of metabolites. Affect the normal growth of seeds...

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Abstract

The invention provides a new application of the rice orphan gene Oscpy and its encoded protein. The present invention finds that the down-regulation of Oscpy expression can promote the germination of rice seeds under flooding conditions and the rapid growth of rice seedlings under normal conditions. Utilizing the functional characteristics of the gene, high-efficiency direct-seeding rice materials can be created, and the gene has great application potential in rice breeding. Oscpy expression down-regulates the rapid growth of rice seedlings, which can improve the efficiency of rice direct seeding, and is also conducive to improving the efficiency of seedling raising in the Yangtze River Basin when it encounters "cold spring".

Description

technical field [0001] The invention belongs to the field of biological genetic engineering, and in particular relates to the application of an orphan gene Oscpy in rice and its coded protein in rice seeding. Background technique [0002] Rice is one of the main food crops in my country, how to improve its resistance to stress has always been one of the problems that biologists generally pay attention to. There are two main methods of rice planting: direct seeding and seedling raising and transplanting. The direct seeding method is the oldest and most primitive planting method left by our ancestors. At the stage when the food and clothing problem is the main contradiction, in order to increase the yield per unit area, we can only make full use of local climate resources for fine farming, so seedling raising and transplanting have always been in a dominant position. However, with the development of my country's economy, the field of agricultural production has also changed f...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/82A01H4/00A01H5/00A01H5/10A01H6/46
CPCC07K14/415C12N15/8218C12N15/8261A01H4/00Y02P60/20
Inventor 彭晓珏阳菁丁霞朱友林胡赵应素平
Owner NANCHANG UNIV
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