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Application of rice transcription factor Os06g08400 genes

A technology of rice transcription factor and gene, applied in the field of genetic engineering

Active Publication Date: 2012-12-12
INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In conclusion, the control of grain development in rice is a complex process involving the coordinated control of multiple genes and multiple pathways. So far, few genes have been found to regulate this trait, and the molecular mechanism of regulating grain development has not yet been elucidated. Therefore, it is necessary to search for genes that control the development of grain traits. and new methods of excavation are of great significance to crop improvement and increase crop yield

Method used

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  • Application of rice transcription factor Os06g08400 genes
  • Application of rice transcription factor Os06g08400 genes
  • Application of rice transcription factor Os06g08400 genes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1 Acquisition of rice transcription factor Os06g08400 gene and construction of plant expression vector

[0026]Find the rice transcription factor OsOrphan40 gene in the Plant Transcription Factor Database (http: / / plntfdb.bio.uni-potsdam.de / v3.0 / fam_mem.php?family_id=Orphans&sp_id=OSAJ), or in the MSU Rice Genome Annotation Project Database Search for the number Os06g08400, and design PCR amplification primers according to its CDS sequence: forward primer F1: 5′-CAAAAAAGCAGGCTTCATGGGTTGGTTGACCAAATTTTTTAGA-3′ and reverse primer R1: 5′-CAAGAAAGCTGGGTCAAATGGGAAAGTCCCTGTTAACCG-3′.

[0027] Using rice 'Nipponbare' (Oryza sativa subsp. japonica) leaves as materials, total RNA was extracted by TRIzol method, and cDNA was obtained by reverse transcription. The reaction steps were as follows: (1) On ice, add The following materials: total RNA 6 μl (0.1ng-5 μg), Oligo(dT)18 primer 1 μl, nuclease-free ddH 2 O5 μl, placed in a PCR instrument at 65°C for 5 minutes; (2) Then ...

Embodiment 2

[0032] Embodiment 2 Obtaining of transgenic rice plants

[0033] Take the mature seeds of rice 'kitaake', shell them manually or mechanically, select the plump, smooth and spot-free seeds and inoculate them on the induction medium for induction culture after being sterilized. Select the rice callus with good appearance and good growth as the recipient material, and use the Agrobacterium-mediated method to transfer the vector ubi:VP64-OsOrphan40 into the rice callus, and use acetosyringone containing 100μM and an O.D. value of 0.7 The AAM transformation solution of Agrobacterium was transformed, and the callus soaked in the transformation solution was placed on the co-culture medium for co-cultivation, cultured in the dark at 25°C for 3 days, then placed on the screening medium for about 30 days, and subcultured once every 10 days . Then, the screened out resistant calli were transferred to the differentiation medium for differentiation for about 20 days, and subcultured every...

Embodiment 3

[0038] Example 3 Identification of Os06g08400-VP64 transgenic rice

[0039] In order to detect whether the CDS sequence of the Os06g08400 gene is integrated into the rice genomic DNA, the total DNA of wild-type and transgenic rice leaves were extracted by CTAB method, and the detection primer sequences were designed. The upstream primer: 5′-ATTTTTTTAGCCCTGCCTTCATACG-3′, the downstream primer: 5′ '-CCAAATGTTTGAACGATCGATCCA-3', for PCR, the amplification system is: reaction buffer 5μl, dNTP 1μl, ddH 2 O 2.2 μl, Taq DNA polymerase 0.2 μl, upstream and downstream primers 0.3 μl each, DNA template 1 μl, the reaction conditions were hot start at 96°C for 5 minutes; 95°C for 30s, 55°C for 30s, 72°C for 2min, a total of 32 cycles; 72°C Extend for 10 minutes, and finally finish the reaction at 25°C. The transgenic plants were identified, and the amplified products were subjected to agarose gel electrophoresis. The electrophoresis results showed that the target bands were amplified in ...

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Abstract

The invention relates to an application of rice transcription factor Os06g08400 genes. A transcription factor activation motif VP64, namely four transcription factor activation motifs VP16, is combined with the rice transcription factor Os06g08400 genes to establish a combined transcription factor, and then the combined transcription factor is converted into crops such as rice so that the characters of rice grains are improved, for example, the rice grains are obviously widened and thickened. The application related by the invention has an important theoretical value for detail definitions of a mechanism for regulating and controlling seed development, and the characters of the rice grains can be improved through a transgene means, so that the application related by the invention is also of importance in production practices.

Description

technical field [0001] The invention belongs to the field of genetic engineering, and in particular relates to the application of rice transcription factor Os06g08400 gene. Background technique [0002] Rice (Oryza sativa L.) is one of the three most important food crops and is the staple food of more than half of the world's population. The current research on increasing rice yield is more dependent on limited rice germplasm resources, the advantages of traditional hybrid breeding are gradually weakening, and rice transgenic technology may explore the potential of further increasing rice yield. [0003] The yield of cereal crops largely depends on its grain size, so gene regulation of rice grain traits is an important research area. In recent years, at least 8 genes have been revealed to play an important role in controlling grain shape through gene cloning, QTL and other technical means. The SG1 gene encodes a protein of unknown function, which is mainly expressed in ric...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/63C12N5/10C12N1/21C12N15/82A01H5/00C12N15/11
Inventor 赵涛李涛李宏宇刘斌刘军林辰涛
Owner INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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