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A Rhizobium wys3r1 Strain of Cassia and Its Application

A technology of Cassia and bradyrhizobium, applied in the field of microorganisms, can solve the problems of low nitrogen fixation efficiency, affecting planting and application, slow nodulation, etc., and achieves strong nitrogen fixation ability, promotion of fertilization and soil fertility, and high nodulation rate Effect

Inactive Publication Date: 2020-11-17
FUJIAN AGRI & FORESTRY UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, under natural conditions, Cassia rotifera forms nodules slowly and rarely, and its nitrogen fixation efficiency is low, which seriously affects its planting and popularization and application.

Method used

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  • A Rhizobium wys3r1 Strain of Cassia and Its Application
  • A Rhizobium wys3r1 Strain of Cassia and Its Application
  • A Rhizobium wys3r1 Strain of Cassia and Its Application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 Isolation and Purification of Rhizobium WYS3R1

[0045] 1 Isolation of rhizobia

[0046] Take fresh, mature and large nodules of Cassia rotifera, rinse them with water, and absorb the surface moisture with filter paper. Put it in 95% (w / v) ethanol for 3-5 minutes, take it out and rinse it with sterile water for 5-6 times, then put it in 0.1% (w / v) HgCl 2 Sterilize for 3-5 minutes, take it out and rinse with sterile water 5-6 times. Then cut in half on a flame-sterilized glass slide, clamp half of the tumor with sterile tweezers, make the incision face the surface of YMA (Table 1) culture medium, and incubate at 28°C after being upside down.

[0047] 2 purification

[0048] After the growth of bacteria, a small amount of rhizobia colonies were scraped from the plate with the tip of a pipette, diluted with 1 mL of sterile water, and then cultured by streaking on the plate again. After 3 days, the colonies were observed until 15 days (slow It takes 7-15 days ...

Embodiment 2

[0054] Example 2 Identification of 16S rDNA Molecular Biology of Rhizobium WYS3R1

[0055] Perform PCR-specific amplification of 16S rDNA on rhizobia monoclonal liquid, and the forward primer is SEQ ID NO.3: V4V5515 -F 5'-GTGCCAGCMGCCGCGGTA-3'; the reverse primer is SEQ ID NO.4: V4V5907 -R 5'-CCGTCAATTCCTTTGAGTTT-3'. The enzyme is 2×star Mix. The PCR amplification products were detected by electrophoresis imaging technology to observe whether they had bands, and the remaining PCR amplification products were used for sequence determination. The sequencing result is shown in SEQ ID NO:5. The PCR reaction system is shown in Table 3.

[0056] Reaction conditions: 95°C for 5 min; (95°C for 20 s, 55°C for 20 s, 72°C for 50 s) × 44 cycle; 70°C for 5 min.

[0057] Obtain 13 reference strain sequences from the NCBI (GenBank) database, use the software BioEdit and MEGA6 to analyze the 16S rDNA partial sequences of the isolated strains and the reference strains, and construct the ...

Embodiment 3

[0058] Example 3 Tieback test

[0059] The purpose of the experiment: to screen out the rhizobia with high binding efficiency and strong nitrogen fixation ability with Cassia forage.

[0060] Test materials: Test plant: Minyu No. 1 Cassia rotifera; Test strain: isolated and purified Rhizobium.

[0061] Main test instruments and equipment: artificial climate growth chamber, ultra-clean workbench, high-pressure sterilization pot, constant temperature incubator, 25 potted pots of 15×15 cm, 2 beakers of 1 L, tweezers, Petri dishes, filter paper, glass Sticks, scissors and gauze etc.

[0062] Test Drugs and Reagents

[0063] (1) Test drugs: mannitol, MgSO 4 ∙7H 2 O, NaCl, yeast powder, K 2 HPO 4 、KH 2 PO 4 , CaCO 3 , Ca(NO 3 ) 2 ∙4H 2 O, MgSO 4 ·7H 2 O, CaCl 2 2H 2 O, Na 2 HPO 4 12H 2 O, C 10 h 12 N 2 o 8 FeNa·3H 2 O, Na 2 MoO 4 , MnSO 4 、H 3 BO 3 、CuSO 4 ·5H 2 O and ZnSO 4 ·7H 2 O.

[0064] Test reagent:

[0065] 1) YMA (Yeast Mannitol Agar) liqu...

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Abstract

The invention discloses a rhizobium strain WYS3R1 and an application thereof, belonging to the technical field of microorganisms. The strain was isolated and purified from fresh nodules of the genus Cassia forage, detected by PCR nodA gene, and identified by 16S rDNA molecular biology, identified as Bradyrhizobium ( Bradyrhizobium ) of the new strain and named Wuyuan 1. It was deposited in the China Center for Type Culture Collection on April 10, 2018, and the deposit number is CCTCC NO: M2018190. The rhizobium WYS3R1 of the present invention has the characteristics of efficient nodulation and strong nitrogen fixation ability through the laboratory pot plant back-connection test, and inoculation of the rhizobium can significantly increase the number of nodules, the nitrogen-fixing efficiency of nodules, the biomass and plant nitrogen of the forage of the genus Cassia content, and then achieve the effect of fertilizing soil fertility and improving the ecological environment.

Description

technical field [0001] The invention belongs to the field of microorganisms, and in particular relates to a Cassia Rhizobium WYS3R1 strain and its application. Background technique [0002] my country has about 56.9 million hm 2 Acidic soil accounts for more than 42.2% of the total cultivated land area. In recent years, with the application of large amounts of chemical nitrogen fertilizers, soil acidification has become increasingly serious. The acidic soil in my country is mainly concentrated in the area south of the Yangtze River. This area is rich in light, heat, water and other resources, but the soil is thin, sticky, low in pH and nutrient availability, which seriously restricts the rapid development of agriculture in this area. In terms of production, crop yields are often increased by applying large amounts of chemical fertilizers. However, the input of a large amount of chemical fertilizers will not only lead to soil compaction and further acidification, but also ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/20C05F11/08A01N63/20A01P21/00C12R1/01
CPCA01N63/00C05F11/08C12N1/205C12R2001/01
Inventor 廖红杨庆李欣欣
Owner FUJIAN AGRI & FORESTRY UNIV
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