Application of extremely halophilic archaea NaSOD gene in improving rice salt tolerance

A technology of extreme halophilic archaea and salt tolerance, which is applied in the application field of extreme halophilic archaea NaSOD gene in improving the salt tolerance of rice, and can solve problems such as yield reduction

Inactive Publication Date: 2012-04-04
CHINA JILIANG UNIV +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, due to the existence of a large amount of salinized soil, a considerable part of

Method used

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  • Application of extremely halophilic archaea NaSOD gene in improving rice salt tolerance
  • Application of extremely halophilic archaea NaSOD gene in improving rice salt tolerance
  • Application of extremely halophilic archaea NaSOD gene in improving rice salt tolerance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1 Construction of expression vector p1301-NaSOD

[0024] Cultivate Escherichia coli containing the plasmid pGEX-SOD (patent application 200810163748.8), use the specific primers of the NaSOD gene of the halophilic archaea (respectively adding Pst I and Kpn I restriction sites) to carry out bacterial liquid PCR, and recover the purified PCR product Connected to the intermediate vector pMD19-T, transformed into Escherichia coli DH5α. Positive clones were picked and identified by PCR and sequencing. Shake the correct bacterial solution to extract the plasmid pMD19-NaSOD. The plasmid pMD19-NaSOD was double-digested with Pst I and Kpn I, then inserted into the multiple cloning site of pCAMBIA1301, and the CaMV35S promoter and NOS terminator were respectively inserted before and after the target gene to obtain the plant expression vector p1301-NaSOD (such as figure 1 shown). After the recombinant was identified by enzyme digestion, it was introduced into Agrobacter...

Embodiment 2

[0028] Example 2 Genetic transformation of rice mediated by Agrobacterium

[0029] The mature embryos of rice were used as explants to induce and culture the callus, and the embryogenic callus was selected as the transformation recipient. According to Hiei et al. (Hiei Y, Ohta S, Komari T, Kumashiro T. Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA [J]. Plant. 1994, 6: 271-82.), through transforming Agrobacterium tumefaciens EHA105 carrying the plant expression vector p1301-NaMnSOD into rice callus tissue, and through a series of screening, transgenic rice was differentiated. details as follows:

[0030] (1) Induction and subculture of rice mature embryo callus: After the mature seeds of Oryza sativa L.cv.Nipponbare were dehulled, they were disinfected with 70% alcohol for 2 minutes, and 20% (V / V) sodium hypochlorite solution (plus 2 ~3 drops of Tween-200) soaked for 30min, washed the seeds wi...

Embodiment 3

[0036] Example 3 Identification of transgenic plants

[0037] Hygromycin screening: After the regenerated seedlings are transplanted into the greenhouse, select new leaves in the same position, take 2-3 cm, and soak them in sterile water containing 50 mg / L hygromycin and 1.0 mg / L 6-BA. After 5 days, the leaves of non-transgenic rice plants turned yellow and brown, the leaves of false positive plants also turned yellow, and the leaves of transformed plants remained green.

[0038] PCR detection of transgenic rice: the total DNA of rice leaves was extracted by SDS method, amplified with NaSOD specific primers, and all 14 transgenic samples had a band at 621bp (such as figure 2 shown).

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Abstract

The invention discloses an application of an extremely halophilic archaea (Natrinema altunense sp.) NaSOD gene in improving rice salt tolerance. The base sequence of the extremely halophilic archaea NaSOD is shown by SEQ ID NO.3. The over-expression NaSOD genetically modified rice has the advantages of higher ROS scavenging capability and capability of reducing membrane lipid peroxidation caused by salt treatment so that higher photosynthetic rate can be guaranteed. According to the invention, the extremely halophilic archaea NaSOD gene is transferred into the rice so that the salt tolerance of the rice can be obviously improved.

Description

technical field [0001] The invention relates to the field of functional genomics, in particular to the application of NaSOD gene of extreme halophilic archaea in improving the salt tolerance of rice. Background technique [0002] With the deterioration of the ecological environment, the development of facility agriculture and the improper use of chemical fertilizers, soil salinization has become a worldwide resource problem and ecological problem, and the area of ​​saline-alkali land is increasing year by year (Mahajan S, Tuteja N.Cold, salinity and drought stresses: an overview [J]. Arch.). According to incomplete statistics, the saline-alkali land area in the world is about 954 million hm 2 (FAO, 2008). my country has a large area of ​​saline-alkali soil, widely distributed, and various types. The total area of ​​various types of saline-alkali land is 99.133 million hm 2 , of which the modern saline-alkali soil area is 36.93 million hm 2 , about 44.87 million hm of res...

Claims

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Application Information

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IPC IPC(8): C12N15/82C12N15/53A01H5/00
Inventor 朱诚陈珍杨卫军潘艳会
Owner CHINA JILIANG UNIV
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