Cassia rhizobium strain TXN1 and application thereof

A technology of TXN1 and Cassia, which is applied in the field of microorganisms, can solve the problems of affecting planting and popularization, low nitrogen fixation efficiency, and slow nodulation, etc., and achieves the effects of strong nitrogen fixation ability, increased total nitrogen content, and high nodulation rate

Inactive Publication Date: 2019-02-01
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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  • Cassia rhizobium strain TXN1 and application thereof
  • Cassia rhizobium strain TXN1 and application thereof
  • Cassia rhizobium strain TXN1 and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 Isolation and Purification of Rhizobium TXN1

[0045] 1 Isolation of rhizobia

[0046] Take fresh, mature and large nodules of Cassia pinnata, rinse them with water, and dry 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 Rhizobia need 7-15 days to app...

Embodiment 2

[0054] Example 2 Identification of 16S rDNA Molecular Biology of Rhizobium TXN1

[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 416S 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 of...

Embodiment 3

[0060] Example 3 Tieback test

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

[0062] Test materials: Test plant: Minyu No. 1 Cassia round leaves; Test strain: isolated and purified nitrogen-fixing bacteria 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] ...

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Abstract

The invention discloses a cassia rhizobium strain TXN1 and application thereof and belongs to the technical field of microorganism. The cassia rhizobium strain TXN1 is obtained by being separated andpurified from fresh nodules of cassia grasses and performing PCR (polymerase chain reaction) detection on nodA (nodulation protein A) gene as well as 16S rDNA (ribosomal deoxyribonucleic acid identification) molecular biological detection, determined to be a new strain of Bradyrhizobium and named as Tianyu-1. The cassia rhizobium strain TXN1 is preserved in China Center for Type Culture Collectionin April 10, 2018 with a preservation number of CCTCC NO: M2018191. Laboratory pot culture inoculation experiments prove that the cassia rhizobium strain TXN1 has the advantages of efficient nodulation and large nitrogen fixing capacity; inoculation of the cassia rhizobium strain TXN1 can significantly increase the number of root nodules, the root nitrogen fixing efficiency, the biomass and the plant nitrogen content of cassia grasses and further achieve the effects of increasing 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 TXN1 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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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20A01N63/00A01P21/00C12R1/01
CPCA01N63/00C12N1/205C12R2001/01
Inventor 廖红杨庆李欣欣
Owner FUJIAN AGRI & FORESTRY UNIV
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