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Chamaecrista rhizobium TXR2 and application thereof

A technology of TXR2 and Cassia genus, applied in Cassia Rhizobium TXR2 and its application fields, can solve problems affecting planting and popularization and application, slow nodulation, low nitrogen fixation efficiency, etc., and achieve strong nitrogen fixation ability and high nodulation rate , the effect of increasing the total nitrogen content

Inactive Publication Date: 2019-02-15
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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  • Chamaecrista rhizobium TXR2 and application thereof
  • Chamaecrista rhizobium TXR2 and application thereof
  • Chamaecrista rhizobium TXR2 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 Isolation and purification of Azotobacter tumefaciens strain TXR2

[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 streaked on the plate for culture again. After 3 days, the colonies were observed until 15 days (slow Rhizo...

Embodiment 2

[0054] Example 2 Identification of 16S rDNA Molecular Biology of Rhizobium TXR2

[0055] Perform PCR-specific amplification of 16S rDNA on rhizobia monoclonal liquid, and the forward primer is SEQ ID NO.3: V4V5515 -F 5'-GTGCCAGCMGCCGCGGTAA-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 4.

[0056] Table 4 16S rDNA2×starMix enzyme reaction system

[0057]

[0058] 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.

[0059] Obtain 13 reference strain sequences from the NCBI (GenBank) database, use the software BioEdit and MEGA6 to analyze the 16S rDNA partial sequences o...

Embodiment 3

[0060] Example 3 Tieback test

[0061] The purpose of the experiment: to screen out the rhizobia with high binding efficiency and strong nitrogen fixation ability with Cassia forage.

[0062] Test materials: Test plant: Minyu No. 1 Cassia rotifera; Test strain: isolated and purified Rhizobium.

[0063] 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.

[0064] Test Drugs and Reagents

[0065] (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.

[0066] Test reagent:

[0067] 1) YMA (Yeast Mannitol Agar) liqu...

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Abstract

The invention discloses rhizobium TXR2 and the application thereof, and belongs to the technical field of microbes. The strain is isolated and purified from fresh root nodules of chamaecrista pasture;through PCR detection of a nodA gene as well as 16S rDNA molecular biological identification, the rhizobium TXR2 is determined to be a novel strain of bradyrhizobium, and is named as Tianyuan No.2. The rhizobium TXR2 is collected in China Center for Type Culture Collection on April 10, 2018, with the collection number of CCTCC NO: M2018193. A pot culture tie-back test in a laboratory proves thatthe rhizobium TXR2 has the characteristics of high-efficiency nodulation and strong nitrogen-fixing ability; inoculation with the rhizobium can significantly increase the number of the root nodules ofthe chamaecrista pasture, the nitrogen nitrogen-fixing efficiency of the root nodules, the biomass and the nitrogen content of plants, and the effects of improving the soil fertility and improving the ecological environment are achieved.

Description

technical field [0001] The invention belongs to the field of microorganisms, and in particular relates to a Cassia Rhizobium TXR2 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 cause eut...

Claims

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Application Information

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IPC IPC(8): C12N1/20C05F11/08C12R1/41
CPCC05F11/08C12N1/205C12R2001/41
Inventor 廖红杨庆李欣欣
Owner FUJIAN AGRI & FORESTRY UNIV
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