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Application of synthetic transcription factor VP64-Os01g63510

A rice transcription factor, VP16 technology, applied in the fields of application, hybrid peptides, angiosperms/flowering plants, etc., can solve the problem of no discovery of WUSCHEL family transcription factors regulating grain size, etc., achieve good market application value, and increase the effect of thousand-grain weight

Active Publication Date: 2013-07-31
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

Although many of the above transcription factor families are involved in the regulation of rice grain size, no research has been found on the regulation of grain size by WUSCHEL family transcription factors

Method used

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  • Application of synthetic transcription factor VP64-Os01g63510
  • Application of synthetic transcription factor VP64-Os01g63510
  • Application of synthetic transcription factor VP64-Os01g63510

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1 Isolation of Os01g63510 gene and construction of plant expression vector

[0031] The Os01g63510 gene was found in the plant transcription factor database (http: / / planttfdb.cbi.edu.cn / index.php?sp=Osj), and PCR amplification primers (F1: 5'-CAAAAAAGCAGGCTTCATGGAGGCTCTTAGCGGG-3' and Reverse primer

[0032] R1: 5'-CAAGAAAGCTGGGTC GAGGCCGAAGCTGCA-3') Using the total cDNA of wild Nipponbare rice as a template, PCR was performed to obtain the CDS sequence of the Os01g63510 gene. The nucleotide sequence is shown in SEQ ID NO.1.

[0033] Perform PCR according to PrimeSTAR polymerase amplification system and reaction program. This process consists of two rounds of PCR. The primers of the first round of PCR use the gene primers (F1 and R1) with some adaptor attB adapters, and the template of the second round uses the PCR products of the first round, and the primers use the complete adaptor attB Primer [attB5'adaptor:

[0034] 5'-GTGGGGACAAGTTTGTACAAAAAAGCAGGCTTC-3', attB3'

...

Embodiment 2

[0036] Example 2 Obtaining Transgenic Rice Plants

[0037] Take the mature seeds of rice ‘kitaake’, manually or mechanically dehull them, select the seeds with full and clean aseptic spots, sterilize them, and inoculate them on the induction medium for induction culture. Select rice callus with good appearance and good growth ability as the acceptor material, use Agrobacterium-mediated method to transfer ubi:VP64-Os01g63510 into rice callus, and use 100μM acetosyringone and OD value 0.7 The AAM transformation solution of Agrobacterium was transformed. The callus soaked in the transformation solution was placed on a co-culture medium for co-cultivation, cultured in the dark at 25°C for 3 days and then cultured on a screening medium for about 30 days, and subcultured once every 10 days. Then transfer the screened resistant callus to differentiation medium for about 20 days, and subculture once every 10 days. Transfer the resistant callus from the green seedlings to a rooting mediu...

Embodiment 3

[0042] Example 3 Identification of Transgenic Rice

[0043] In order to detect whether the Os01g63510 gene is integrated into the rice genomic DNA, wild-type ('kitaake') and transgenic rice leaf DNA prepared by the present invention were extracted, and a pair of primers (VP64-1F: 5'-) were designed from the plant expression vector at both ends of the gene. ATGGACGCGCTGGACGATTT-3', 2424-R: 5'GAGGCCGAAGCTGCAAA3') PCR was carried out to identify transgenic plants according to PrimeSTAR polymerase amplification system and reaction program. The PCR results showed that the target band (867bp) was amplified in all transgenic rice, while the wild There is no band for this purpose in type rice, the result is image 3 . It is preliminarily determined that the Os01g63510 gene has been inserted into the rice genome by Agrobacterium-mediated method, and the PCR product was analyzed by sequencing results to be the target gene sequence.

[0044] In order to detect the overexpression of the ubi:V...

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Abstract

The invention provides application of a synthetic transcription factor VP64-Os01g63510. Transcription factor activation motifs VP64 (namely the four transcription factor activation motifs VP16) and paddy rice transcription factors Os01g63510 are constructed to obtain the synthetic transcription factor in a gene fusion mode. The synthetic transcription factor VP64-Os01g63510 is transformed into paddy rice to improve characters of paddy rice seeds. Therefore, the genetically modified paddy rice seeds are obviously lengthened and widened, the thousand seed weight is obviously increased, and the market application prospects are good.

Description

Technical field [0001] The invention belongs to the field of genetic engineering, and specifically relates to the application of the rice transcription factor Os01g63510 gene and the application of the synthetic transcription factor VP64-Os01g63510. Background technique [0002] Rice (Oryza sativa L.) is an important food crop in my country and the world. More than half of the world's population feed on rice as the staple food. Food security has always been a concern around the world, so the yield traits of rice have always been paid attention to by scientists, and rice grain size is one of the main factors affecting yield. At the same time, rice is a model plant for the study of functional genes of crops, and its related genetics and molecular biology research has been paid much attention by researchers. The growth and development of rice grains is an orderly and selective gene expression process, transcription factors It plays a key role in the precise regulation of genes. [0...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/82C12N5/10C12N1/21A01H5/00
Inventor 刘军邹小花李宏宇赵涛刘斌林辰涛
Owner INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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