Rice root system bacillus cereus SaltT1 and application thereof

Abstract

The invention discloses bacillus cereus SaltT1 for rice root systems and application of the bacillus cereus SaltT1, and relates to the technical field of biopesticides, and the technical key points are as follows: the bacillus cereus SaltT1 is used for improving the salt tolerance of rice; the biological source is mainly used for relieving salt stress in the germination period of rice seeds and enhancing salt tolerance in the germination period of the rice seeds, the biological source is environmentally friendly and non-toxic, the biological source is used for soaking the rice seeds for saline-alkali soil planting, the influence on the ecological environment is small, and the biological source is safe and reliable for eating of rice and processed products of the rice; the culture condition requirement is low, and the development and application prospects are good.

Description

Technical Field

[0001] The invention relates to the technical field of biological pesticides, and in particular to a rice root system Bacillus cereus Salt T1 and an application thereof. Background Art

[0002] Rice (Oryza sativa L.) is one of the main food crops for more than half of the world's population, and its output is an important guarantee for world food security. With the growing population, the improvement of rice yield and quality faces increasingly severe challenges. However, during the growth process of rice, it is often subject to various biological stresses (diseases, insects) and abiotic stresses (high / low temperature, salinity, drought / waterlogging, etc.). Among them, high salt stress is one of the abiotic stresses that has the greatest impact on rice growth and development. What is more serious is that there are 1 billion hm2 of saline-alkali lands worldwide, and the area of ​​saline-alkali lands in my country is as high as 90 million hm2, and is further expanding. Therefore, the rational use of saline-alkali land to grow rice has become one of the important measures to ensure the current grain planting area and grain output.

[0003] Studies have shown that physical methods such as irrigation to wash away salt and drainage, and chemical methods such as adding soil conditioners such as gypsum can effectively improve soil physical and chemical properties, but they often cause problems such as water waste or environmental pollution. In addition, although creating and cultivating new salt-tolerant rice germplasms and new varieties is the most economical and effective method, the existing strong salt-tolerant germplasm resources are relatively few, there are not many cloned rice salt-tolerant genes, and the breeding cycle is long, which greatly limits the cultivation of rice on saline-alkali land.

[0004] To this end, the present invention aims to provide a rice root cereus Bacillus cereus SaltT1 and its application to solve the above problems. Summary of the invention

[0005] The invention aims to solve the above problems and provides a rice root cereus Bacillus cereus SaltT1 and an application thereof. The strain is separated from the rice root system of Guanghong No. 3 salt-tolerant rice variety planted in Taishan test base of salt-alkali-tolerant rice research center of Rice Research Institute of Guangdong Academy of Agricultural Sciences in Beidou Town, Taishan City, Guangdong Province, and can be well colonized in rice tissues; it has a significant alleviating effect on salt stress during rice seed germination, improves the salt tolerance of rice seeds during germination, and the biological source is environmentally friendly and non-toxic. It can be used to soak rice seeds to resist salt stress during germination, has little impact on the ecological environment, and is safe and reliable for rice consumption and processed products.

[0006] In order to achieve the above object, the technical solution of the present invention is as follows:

[0007] The present invention provides a rice root cereus Bacillus cereus SaltT1, wherein the Bacillus cereus SaltT1 was deposited in Guangdong Provincial Microbial Culture Collection Center on July 30, 2024, and the deposit address is Guangdong Provincial Institute of Microbiology, 5th Floor, Building 59, Compound No. 100, Xianlie Middle Road, Guangzhou, and the deposit number of the Bacillus cereus SaltT1 is GDMCC No: 64923.

[0008] The invention discloses an application of a rice root cereus Bacillus cereus SaltT1, wherein the Bacillus cereus SaltT1 is used to improve the salt tolerance of rice during the germination period, and the rice variety is the salt-sensitive variety Zhefu 802.

[0009] Bacillus cereus SaltT1 was isolated and purified from rice roots, and the rice root Bacillus cereus SaltT1 was cultured on LB medium at 28-30°C for 48 hours. The colonies were grayish white, opaque, rough, flat, irregular, and like molten wax.

[0010] Bacillus cereus SaltT1 is inoculated into LB liquid culture medium for cultivation, centrifuged, supernatant is removed, and the mixture is resuspended with sterile pure water to obtain a Bacillus cereus biological preparation. The Bacillus cereus bacterial suspension is used for improving salt tolerance of rice during bud stage.

[0011] Compared with the prior art, this solution has the following beneficial effects:

[0012] 1. The Bacillus cereus SaltT1 obtained by the present invention is derived from the root system of the salt-tolerant variety Guanghong No. 3 planted in saline-alkali land, and can be well colonized in rice tissues (or on the surface); in addition, the bacteria screened from the salt-tolerant rice root system of the present invention are used to promote the salt tolerance of rice seeds during germination, which only shows an increase or decrease in the number of certain microorganisms among specific populations, and does not change the categories of microbial populations inside and outside the rice tissues to affect the stability of the populations. In addition, due to the long-term coexistence and mutualistic symbiosis between the bacteria and rice, a series of survival mechanisms similar to the bacteria are formed. After soaking the rice seeds with the bacteria and planting them in saline-alkali land, the bacteria are safer and more reliable for rice yield, consumption and processed products thereof;

[0013] 2. The Bacillus cereus obtained by the present invention has low requirements for culture conditions and has good development and application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] Figure 1It is a single colony image of Bacillus cereus SaltT1 on LB solid medium in the embodiment of the present invention;

[0015] Figure 2 It is a diagram showing the effect of Bacillus cereus SaltT1 and its bacterial suspension on the salt-sensitive rice variety Zhefu 802 during the germination period and a statistical result diagram of the root length and shoot length of seed germination in the embodiments of the present invention. DETAILED DESCRIPTION

[0016] In order to enable those skilled in the art to better understand the solution of the present invention, the technical solution of the present invention will be further described in detail below in conjunction with the embodiments of the present invention and the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work should fall within the scope of protection of the present invention.

[0017] It should be noted that, in the absence of conflict, the embodiments and features in the embodiments of the present invention can be combined with each other. The present invention will be described in detail below in conjunction with the embodiments.

[0018] Example 1: Isolation, purification and preservation of a rice root strain of Bacillus cereus SaltT1, the steps are as follows:

[0019] (1) Preparation of LB solid medium: weigh 10 g of tryptone, 5 g of yeast extract, and 10 g of sodium chloride, add 1000 mL of water and stir evenly. When preparing LB solid medium, add 20 g of agar, heat thoroughly to dissolve, and then divide into portions. Sterilize at 121°C for 20 min and store for later use.

[0020] (2) Isolation and purification of Bacillus cereus SaltT1 from rice roots:

[0021] The rice roots used for root microorganism isolation were collected from the salt-tolerant rice variety Guanghong No. 3 at the Taishan Experimental Base of the Salt-Alkali Tolerant Rice Research Center, Rice Research Institute, Guangdong Academy of Agricultural Sciences, Beidou Town, Taishan City, Guangdong Province.

[0022] The specific operation of the root bacteria isolation method is as follows: weigh 10g of Guanghong No. 3 roots (including soil) into a pre-sterilized conical flask, add an appropriate amount of sterile pure water (just enough to immerse the root tissue and soil), seal the bottle and shake it at 150rpm for 5 minutes, finally place it in a clean workbench and let it stand for 15 minutes, aspirate 100μL of the upper root extract and apply it to the LB plate, set up 3 replicates for the experimental treatment, and observe the results after culturing at 28℃ for 48 to 72 hours.

[0023] The 16S rRNA sequence of the strain isolated from the rice root system of the Taishan Experimental Base of the Salt-Alkali Tolerant Rice Research Center of the Rice Research Institute of Guangdong Academy of Agricultural Sciences has a homology of 99.78% with the 16S rRNA sequence of the Bacillus cereus HBUAS52549 strain with the accession number OM967117.1 in GenBank. Combined with the colony morphology characteristics of the bacteria on the LB solid medium plate and related physiological and biochemical characteristics tests, it is confirmed that it is classified as Bacillus cereus and named Bacillus cereus SaltT1. The sequence fragment is as follows:

[0024]

[0025] Bacillus cereus SaltT1 was cultured on LB solid medium at 28°C for 48 hours. The colonies were grayish white, opaque, rough, flat, irregular, and like molten wax. Figure 1 shown.

[0026] (3) Deposit of Bacillus cereus SaltT1: The strain was deposited in Guangdong Microbiological Culture Collection Center on July 30, 2024, with the deposit number GDMCC No: 64923.

[0027] Example 2: Determination of the salt tolerance of rice seeds during germination promoted by Bacillus cereus SaltT1

[0028] (1) Preparation of LB liquid medium was the same as in Example 1; except that agar was not added, the preparation of LB liquid medium was the same as that of LB solid medium. The weighed agent was fully dissolved and dispensed into conical flasks (100 mL of culture solution per flask), stoppered, bandaged, sterilized at 121° C. for 20 min, and stored for later use after cooling.

[0029] (2) Preparation of single colonies of Bacillus cereus: Bacillus cereus was inoculated into LB liquid culture medium and cultured with shaking at 28°C for 20 h. The fermentation liquid was streaked onto an LB solid culture medium plate using a sterile inoculation loop for 48 h. A single colony was then picked for streaking and preservation.

[0030] (3) Preparation of Bacillus cereus biological preparation: a single colony of Bacillus cereus prepared in step (2) is inoculated into a conical flask of LB liquid culture medium prepared in step (1), and cultured at 28° C. and a shaking rate of 200 rpm for 20 h to obtain a Bacillus cereus fermentation liquid; the obtained fermentation liquid is centrifuged at 12000 rpm for 10 min, the supernatant is discarded, and the bacteria are resuspended in sterile pure water to obtain a Bacillus cereus bacterial suspension.

[0031] (4) Determination of the effect of Bacillus cereus on the salt tolerance of rice seeds during germination: Take an equal amount of the Bacillus cereus suspension prepared in step (3), add an appropriate amount of NaCl to make the NaCl concentration of the suspension 0.7%, use sterile pure water and 0.7% NaCl solution as controls, treat the seeds of the salt-sensitive rice variety Zhefu 802, and conduct a germination test under the same temperature and light conditions. Count the length of the germinated roots and shoots, and take pictures. Figure 2 shown.

[0032] The above specific embodiments are merely explanations of the present invention and are not limitations of the present invention. After reading this specification, those skilled in the art may make modifications to the embodiments without any creative contribution as needed. However, such modifications are protected by the patent law as long as they are within the scope of the claims of the present invention.

Claims

1. A rice root Bacillus cereus SaltT1, Its characteristics are: The deposit number of the Bacillus cereus SaltT1 is GDMCC No: 64923, and the deposit address is: Guangdong Institute of Microbiology, Building 5, Building 59, Dayuan, No. 100 Xianlie Middle Road, Guangzhou; Bacillus cereus SaltT1 sequence fragment is: CCCACCGACTTCGGGTGTTACAAACTCTCGTGGTGTGACGGGCGTGTGTACAAGGCCCGGGAACGTATTCACCGCGGCATGCTGATCCGCGATTACTAGCGATTCCAGCTTCATGTAGGCGAGTTGCAGCCTACAATCCGAACTGAAAACGGTTTATGAGA TTAGCTCCACCTCGCGGTCTTGCAGCTCTTTGTACCGTCCATTGTAGCACGTGTGTAGCCCAGGTCATAAGGGGCATGATGATTTGACGTCATCCCCACCTTCCTCCGGTTGTCACCGGCAGTCACCTTAGAGTGCCCAACTTAATGATGGCAACTAAGATCAAGGGTTGCGCTC GTTGCGGGACTTAACCCAACATCTCACGACACGAGCTGACGACAACCATGCACCACCTGTCACTCTGCTCCCCGAAGGAGAAGCCCTATCTCTTAGGGTTTTCAGAGGATGTCAAGACCTGGTAAGGTTCTTCGCGTTGCTTCGAATTAAACCAATGCTCCACCGCTTGTGCGGCC CCCGTCAATTCCTTTGAGTTCAGCCTTGCGCCGTACTCCCCCAGGCGGAGTGCTTAATGCGTTAACTTCAGCACTAAAGGGCGGAAACCCTCTAACACTTAGCACTCATCGTTTACGGCGTGGACTACCAGGGGTATCTAATCCTGTTGCTCCCCACGCTTTCGCGCCTCAGT GTCAGTTACAGACCAGAAAAGTCGCTTCGCCACTGGTGTTCCTCCATATCTCTCTACGCATTTCACCGCTACACATGGAATTCCACTTTCCTCTCTTCTGCACTCAAGTCTCCCCAGTTTCCAATGACCTCACGGTTGAGCCGTGGGCTTTCACATCAGACTTAAGAAACCACCTGCGC GCGCTTTACGCCCAATAATTCCGGATAACGCTTGCCACCTACGTATTACCGCGGCTGCTGGCACGTAGTTAGCCGTGGCTTTCTGGTTAGGTACCGTCAAGGTGCCAGCTTATTCAACTAGCACTTGTTCTTCCCTAACACAGAGTTTTACGACCCGAAAGCCTTCATCACTCAC GCGGCGTTGCTCCGTCAGACTTTCGTCCATTGCGGAAGATTCCCCTACTGCTGCCTCCCGTAGGAGTCTGGGCCGTGTCTCAGTCCCAGTGTGGCCGATCACCCTCTCAGGTCGGCTACGCATCGTTGCCTTGCCTTGGTGAGCCGTTACCTCACCAACTAGCTAATGCGACGCGGGGTCCAT CCATAAGTGACAGCCGAAGCCGCCTTTCAATTTCGAACCATGCGGTTCAAAATGTTATCCGGTATTAGCCCCGGTTTCCCGGAGTTATCCCAGTCTTATGGGCAGGTTACCACGTGTTACTCACCCGTCCGCCGCTAACTTCATAAGAGCAAGCTCTTAATCCATTCGCTCGAC.

2. The application of a rice root Bacillus cereus SaltT1 as claimed in claim 1, Its characteristics are: The Bacillus cereus SaltT1 is used to improve salt tolerance in rice sprout stage.

3. The application of a rice root Bacillus cereus SaltT1 as claimed in claim 2, Its characteristics are: The rice variety is the salt-sensitive variety Zhehui 802.

4. The application of a rice root Bacillus cereus SaltT1 as claimed in claim 2, Its characteristics are: The salt tolerance detection method of rice sprout stage includes the following steps: S1. Prepare LB liquid culture medium: weigh 10g tryptone, 5g yeast extract, 10g sodium chloride, add 1000mL of water and stir evenly, wait until the above agent is fully dissolved and aliquoted in a conical flask, then plug and bandage, or add 20g / L agar powder to prepare LB solid culture medium, sterilize at 121°C for 20 minutes, cool and store for later use; S2. Preparation of single colonies of Bacillus cereus: Bacillus cereus was inoculated into LB liquid culture medium and cultured in a shaking manner for 20 hours, and then scribble the bacterial liquid and cultured into LB solid culture medium plate for 48 hours, and then single colonies were picked for scribing and preserving; S3. Preparation of Bacillus cereus suspension: Take the single colony of Bacillus cereus and insert it into the conical flask of the LB liquid culture medium, place it at 28°C, and culture it at 200 rpm for 20 hours to obtain the Bacillus cereus fermentation broth; the obtained fermentation broth was centrifuged at 12,000 rpm for 10 minutes, discard the supernatant, and resuspend the bacteria with sterile pure water to obtain the Bacillus cereus suspension; S4. Determination of the promotion effect of Bacillus cereus on salt tolerance during rice germination: Take an equal amount of Bacillus cereus bacteria suspension, add NaCl to make the NaCl concentration of the bacteria suspension reach 0.7%. Use sterile pure water and 0.7% NaCl solution as the control, and treat the seeds of the salt-sensitive rice variety ZheRong 802. Under the same conditions as the temperature and light, a germination test was carried out, and the germination root length and bud length were counted, and photographed and analyzed.

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