| The soil salinization is one of the problems that lead to global land degradation and severely restricts the development of agricultural productivity.With the increase of population and the gradual reduction of cultivated land,the development and utilization of saline-alkali land is of great significance.Bioremediation has the advantages of low input cost,good restoration effect and high ecological benefit.Screening of multifunctional salt-tolerant strains from saline-alkali soils,to explore the interaction in different strain combinations,influence on rice seed germination and related physiological metabolism salt stress,and obtaining the best strain combination for the development of microbial fertilizer.Using microbial fertilizers for rice planting in saline-alkali soils,use RNA-Seq transcriptome sequencing technology to analyze the effects of bio-organic fertilizers on the gene expression of rice leaves in saline-alkali soils,and use orthogonal experiment design to optimize amount of nitrogen fertilizer,phosphate fertilizer,bio-organic fertilizer and microbial agent for rice planting in saline-alkali soils.The main research contents and conclusions are as follows:Four strains of multi-functional saline-alkali tolerant bacteria were identified based on colony morphology,physiological and biochemical characteristics and the homology of 16S rDNA gene sequence.with soil of ningxia saline-alkali land was taken as the material,and SS02 was identified as Bacillus subtilis accession number MW704114,AA04 is Bacillus thuringiensis accession number MW704115,AA05 is Paenibacillus polymyxa accession number MW704116,HS07 is Bacillus aryabhattai accession number MW803060.The strains were resistant to NaCl about 100g/L and alkali tolerant to pH 10,and there was no antagonism between the strains.The nitrogen-fixing capacity of the four strains was 13.25 mg/L,which was the best combination of nitrogen-fixing ability,The increment of available phosphorus in culture medium of different combinations of strains had no correlation with pH value,but was positively correlated with total organic acid content,pH value was negatively correlated with total organic acid content.The yield of total organic acid,IAA and EPS of Bacillus subtilis and Bacillus thuringiensis mixed culture was better than that of other combinations.The AB(Bacillus subtilis+Bacillus thuringiensis)has better interaction than other combinations,and can be used as a potential combination of bacterial strains in microbial fertilizer production.Although four bacterial combinations can also be chosen as one of the options,the fermentation conditions need to be further optimized.Combination of different strains has certain promote effect on seed germination and seedling growth of rice under salt stress,ABD(Bacillus subtilis+Bacillus thuringiensis+Bacillus aryabhattai)and ABCD(Bacillus subtilis+Bacillus thuringiensis+Paenibacillus polymyxa+Bacillus aryabhattai)strain combination can make the germination rate can be increased by 20%or so,,the bud length and root length of rice were increased by 4.75 and 0.72 times,respectively,the vigor index was increased by 3.19 times,and the salt tolerance index was increased by 3.2 times in ABCD combination.The chlorophyll content and root activity were significantly higher than those without salt stress.The content of malondialdehyde(MDA)in rice seedlings under salt stress was decreased,and peroxidase(POD),and catalase(CAT),and superoxide dismutase,(SOD)were increased by adding salt-tolerant bacteria,and they can alleviate the salt toxicity to rice,but there are some differences among different treatments.It has a good application prospect to improve the SOD activity of seedlings by the combination of multiple strains.Application of bio-organic fertilizer can significantly increase rice leaf area and chlorophyll,plant tiller number and dry matter quality(P<0.05),T2(bio-organic fertilizer inactivation +chemical fertilizer)vs T1(bio-organic fertilizer+chemical fertilizer),T3(chemical fertilizer)vs T1(Bio-organic fertilizer+chemical fertilizer),T4(blank control)vsTl(bio-organic fertilizer+chemical fertilizer)and T4(blank control)vs.T3(chemical fertilizer),the number of differential genes in the 4 treatment groups were 6,693,4796,6976,and 1866,respectively,Bio-organic fertilizer combined with chemical fertilizers caused the highest number of differential gene expressions,followed by microorganisms,and single application of chemical fertilizers was the smallest;GO analysis showed that the application of bio-organic fertilizer mainly affects the biosynthesis and metabolism of rice leaf peptides and amides,translation processes,organelles and organelle membranes,etc.;the biological processes of rice leaves was affected by Chemical fertilizers differential genes were not significantly enriched(P>0.05);KEGG analysis showed that the significantly different genes in the application of bio-organic fertilizer were mainly enriched in the ribosome and energy metabolism-related pathways,and ribosome-related genes were expressed more differentially.Possibly there are important regulatory genes for rice to respond to bio-organic fertilizers.The energy metabolism pathway involves photosynthesis and photosynthesis-antenna protein,the photosynthesis-antenna protein metabolism pathway-related genes are significantly up-regulated,and the number of photosynthesis and photosynthesis-antenna protein is up-regulated in T4 vs T1,It is greater than T3 vs T1,T3 vs T1 is greater than T4 vs T3,indicating that the application of bio-organic fertilizer can improve the light-harvesting ability of leaves,enhance photosynthesis,and the effect of bio-organic fertilizer combined with chemical fertilizer is better than that of single application of biological organic fertilizer or chemical fertilizer;T2 vs T1,T3 vs T1,T4 vs T1 treatment of plant hormone signal transduction carotenoid synthesis genes up-regulated,rice stress resistance enhanced;the analysis of salt-tolerant gene transcription factors shows thatbio-organic fertilizer can effectively promote the expression of genes in response to salt stress in plants and enhance the salt tolerance of rice.The growth traits and yield of rice were significant difference under the combined application of four fertilizers(P<0.05),the soil pH,Na+,Cl-and total salt contents were significantly decreased,and the organic matter,alkali-hydrolyzed nitrogen and enzyme activities were increased.The range analysis showed that the increase of nitrogen fertilizer and bio-organic fertilizer made the growth characteristics of rice better.The yield increased with the increase of nitrogen fertilizer and microbial agent,but decreased with the increase of phosphate fertilizer.There was no significant difference between the change of fertilizer amount and soil quality.The comprehensive evaluation showed that N 109.5 kg/hm2,P2O5 150 kg/hm2,biological organic fertilizer 9000 kg/hm2,and microbial agent 90 kg/hm2 could be used as the guiding fertilizer amount for rice planting in moderate saline-alkali land in Ningxia Yellow River Irrigation District. |
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