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Efficient nitrogen-fixing alfalfa rhizobium strain and method for screening molecular marker

A method of alfalfa and molecular markers, applied in the direction of microorganism-based methods, biochemical equipment and methods, bacteria, etc., can solve the problems of inability to achieve high yield, low nitrogen fixation efficiency, mixed types of rhizobia strains, etc., and achieve good affinity , Increased planting yield and good adaptability

Active Publication Date: 2015-05-13
HEBEI UNIVERSITY
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] The purpose of the present invention is to provide a high-efficiency nitrogen-fixing alfalfa rhizobium strain and its molecular marker screening method to solve the problems of mixed types of rhizobia strains obtained by existing screening methods, low nitrogen fixation efficiency, and failure to achieve high yield when applied to alfalfa production. question

Method used

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  • Efficient nitrogen-fixing alfalfa rhizobium strain and method for screening molecular marker
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  • Efficient nitrogen-fixing alfalfa rhizobium strain and method for screening molecular marker

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Experimental program
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Effect test

Embodiment 1

[0025] (1) Sampling and separation and purification: For the alfalfa planted in the surrounding areas of Huanghua, Cangzhou, Hebei, the plants with good growth and robustness were selected, and the root nodules were sampled at the early flowering stage of the alfalfa. At each sampling location, 5-point sampling method was used. Sampling, the sampling area of ​​each point is 1m 2 , each plant selects plump and pink root nodules, selects 3 large root nodules, selects two locations, selects 45 root nodules in each location, and obtains 90 root nodules in two locations;

[0026] To isolate and purify bacterial strains from selected root nodules, the specific steps are:

[0027] (a) Select a complete and undamaged root nodule and fully soak it in sterile water;

[0028] (b) Nodule surface disinfection: 95% ethanol for 3 minutes—0.5% hypochlorous acid for 3 minutes—sterile water rinse 6-8 times;

[0029] (c) Separation by streaking on the plate: crush the sterilized root nodule wi...

Embodiment 2

[0061] Identification of embodiment 2 strains

[0062] The preservation name of the strain provided by the present invention is Sinorhizobium meliloti 729-2-4, the preservation unit is CGMCC, the preservation number is CGMCC No.9755, and the preservation date is October 10, 2014. The address of the depository unit is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Institute of Microbiology, Chinese Academy of Sciences.

[0063] The high-yielding Rhizobium alfalfa strain provided by the present invention has an optimum growth temperature of 28°C and is an aerobic culture. The colony on the YMA medium is translucent, milky white, round, thin at the edge, thick in the middle, and thin. The time is 48-72 hours, the colony size is 3-5mm; the bacterial cell is rod-shaped, with strong refractive hydroxy fatty acid particles, accounting for 1 / 3 of the cell volume, slightly rounded at both ends, and the size is 2-3μm×3- 5 μm, Gram-negative bacteria, Gram stain is red.

Embodiment 3

[0064] The application of embodiment 3 new bacterial strains

[0065]Experimental method: inoculate the bacterial strain obtained in Example 1 of the present invention onto alfalfa planted in the field (around Huanghua), its inoculation method, inoculation area and sampling detection method are shown in step 8 of Example 1; meanwhile, adopt the same The experimental conditions were inoculated with American strain (AM185H2) and Australian strain (AU128) as a comparative experiment.

[0066] Experimental results:

[0067] Table 4 Application results of strains

[0068]

[0069] Note: The Tukey test is used, a b c are different subsets, the above data are the average value of the five-point sampling of the experiment, and three parallel experiments.

[0070] As can be seen from the table, the bacterial strains from Australia and the United States do not have a good effect on improving the yield of alfalfa than the bacterial strains screened by the present invention. It can ...

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Abstract

The invention discloses an efficient nitrogen-fixing alfalfa rhizobium strain. The strain has a collection name of Sinorhizobium meliloti 729-2-4 and is collected in CGMCC on October 10, 2014 and has a collection number of CGMCC No. 9755. The strain is a rhizobium strain which can establish good symbiotic relationship with alfalfa and is strong in nitrogen-fixing capability and the planting yield of the alfalfa can be effectively increased. Simultaneously, the invention also provides a method for screening a molecular marker of the rhizobium strain. The method is scientific, reasonable and simple and easy to operate and the method can be used for more accurately screening the efficient nitrogen-fixing rhizobium strain which is applicable to local climate and soil and establishes good symbiotic relationship with alfalfa.

Description

technical field [0001] The invention relates to a rhizobia strain and a screening method thereof, in particular to a high-efficiency nitrogen-fixing alfalfa rhizobium strain and a molecular marker screening method thereof. Background technique [0002] Medicago sativa, formerly known as: Medicago sativa, also known as: alfalfa, Latin name: Medicago sativa, It is a perennial herbaceous plant belonging to the leguminous family. It has strong roots, penetrates deep into the soil layer, and has a well-developed root neck. It is a kind of forage with good palatability, high nutritional value and development value. It has the title of "King of Forage" in the industry. The growth of alfalfa mainly relies on rhizobia to establish a symbiotic relationship with nitrogen fixation. Studies have shown that the establishment of the symbiotic relationship between rhizobia and alfalfa is the result of the tripartite interaction between bacteria, plants and the environment. Only by establi...

Claims

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Application Information

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IPC IPC(8): C12N1/20C12Q1/68C12Q1/04C12R1/01
CPCC12Q1/6869C12N1/205C12R2001/01
Inventor 刘晓云刘朋飞刘桂霞
Owner HEBEI UNIVERSITY
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