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Rice high-temperature-resistant related gene OsZFP, selection marker and separating method of related gene

Through the screening and positioning technology of polypeptide and gene sequences, the rice high temperature resistance gene OsHTAS is accurately located, which solves the problems of inconsistent data and difficulty in determining candidate genes in the existing technology, improves the tolerance of rice to high temperature, and provides improved rice resistance. Methods of reverse breeding.

Inactive Publication Date: 2014-06-11
ZHEJIANG UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Overexpressing the ZFP177 gene, the transgenic tobacco obtained can tolerate a low temperature of 2°C and a high temperature of 55°C, but is more sensitive to salt stress and drought stress, which shows that ZFP177 plays an important role in various abiotic stresses in plants , but different stresses may have different response mechanisms

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0125] Example 1 Identification of High Temperature Resistance Genes

[0126] In this example, high temperature resistant rice variety HT54 and high temperature sensitive rice variety HT13 (both are rice species indica subspecies ( O. sativa ssp . indica)) is an exemplary test material.

[0127] 1) Cultivation of rice seedlings and setting of high temperature resistance treatment conditions

[0128] (1). Seedling cultivation

[0129] Take the paddy field soil from the same plot, air-dry it naturally, crush and sieve it, divide it into plant growth pots (26cm×18cm×6cm) according to the same weight, add the same amount of water to soak overnight, and then use it for sowing. The test materials were sown after soaking and germination. Each pot is divided into two parts, one half is planted with HT54, the other half is planted with HT13, each variety has 3 rows, and each row has 8 plants.

[0130] (2). High temperature treatment period, treatment temperature and other conditio...

Embodiment 2

[0134] Example 2 Confirmation of new gene markers

[0135] In order to realize the precise positioning of high temperature resistance genes in rice, it is obvious that the SSLP in the prior art is not enough to meet the needs. To this end, the inventors have developed new positioning markers. Specifically, the inventor searched for insertion / deletion polymorphisms (Insertion / Deletion, InDel) through the genome-wide sequence differences between the japonica rice Nipponbare and the indica rice 9311 published by NCBI (http: / / www.ncbi.nlm.nih.gov / ) site. According to the DNA sequence at both ends of this site, use Primer5.0 to design appropriate primers, and use NCBI online Primer blast to ensure the specificity of the primers.

[0136] TA cloning and sequencing analysis

[0137] a) Adding A reaction: Since the purified PCR product is amplified with Proybest enzyme, its end is blunt, and base A needs to be added for TA cloning. The total system for adding A reaction is 20 μL, ...

Embodiment 3

[0146] Example 3 Confirmation of high temperature resistance gene OsHTAS

[0147] Through the aforementioned gene mapping research, the inventors discovered a new gene located on chromosome 9 of the rice genome, namely the dominant high temperature resistant gene OsHTAS . The dominant high temperature gene OsHTAS with recessive allele Oshtas The difference in DNA sequence is only a single nucleotide polymorphism (SNP), which occurs in OsHTAS In the 5'-terminal untranslated region, there are motifs related to high temperature tolerance, salt tolerance and drought tolerance (11T12G), OsHTAS (TTTTTTTTTTTGGGGGGGGGGGG) than Oshtas (TTTTTTTTTTTGGGGGGGGGGG) one more G; dominant high temperature resistance gene (LOC_Os09g15430) OsHTAS The full length of the genome sequence is 4784bp, the full length of the coding sequence (CDS) is 1245bp, and the encoded product is 414aa, which belongs to the zinc finger family cluster protein; rice ( Oryza sativa ) ZFP gene, the spec...

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Abstract

The invention discloses a rice high-temperature-resistant related gene OsZFP, a selection marker and a separating method of the related gene. The invention provides a separated rice endogenous high-temperature-resistant gene (OsZFP gene for short) and a coded polypeptide thereof, a preferred rice cell comprising the high-temperature-resistant gene or the coded polypeptide thereof and a preparation method of a plant cell. The invention further provides a novel method and a novel technology for breeding a high-temperature-resistant new crop variety, including a regulatory sequence relevant to high temperature resistance, a tightly-linked molecular marker for marking the high-temperature-resistant gene and a sequence of the molecular marker.

Description

technical field [0001] The invention relates to the technical fields of plant genetic engineering and molecular breeding. Specifically, it relates to the mapping, isolation, functional analysis, sequence characterization, regulatory sequence identification, and co-segregation molecular marker screening and creation of plant high temperature resistance genes and their application in improving the stress resistance of rice and other crops. A method and program for genetic transformation and molecular marker-assisted selection of a heat-resistant gene in rice. In addition, the present invention also relates to the application of improving the stress resistance of crops such as rice by using the high temperature resistance gene. Background technique [0002] Rice is the most important food crop and the main source of food for more than half of the world's population (Khush, 2005). According to reports, rice production can only meet people's living needs by 2050 at an annual gro...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/63C12N5/10C12N1/19C12N15/11C12Q1/68C12N15/10C12N15/82A01H5/00
CPCC07K14/415A01H5/10C12N15/8271A01H1/04C12Q1/6895C12Q2600/13C12Q2600/156A01H1/045C12Q1/6876
Owner ZHEJIANG UNIV
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