Isolation,Antagonistic Screening And Growth Promoting Research Of Pea Endophytes

Plant endophyte refers to a bacterium that exists in plant healthy tissue and can establish a mutually beneficial symbiotic relationship with the host plant without causing substantial damage to the plant.Therefore,they are also called a huge and valuable resource pool in the plant micro-ecosystem.In order to explore the potential microbial resources of Tibetan peas,this study isolates and screens endophytic bacteria from the roots,stems,leaves,and nodules of pea plants collected in Tibet.IAA ability,the antagonistic effect of the strain was measured by the plate method.Field experiments were conducted on the farm of the Tibet Academy of Agriculture and Animal Husbandry to study the growth-promoting effects of selected strains on pea plant height,root length,fresh weight,dry weight,nodules and chlorophyll.The total nitrogen,total phosphorus and total potassium contents of pea plants were measured,and the effects of the strains on pea plants were investigated.Gram stain,contact enzyme test and methyl red test were used to explore the physiological and biochemical characteristics of the strain.The test results are as follows:(1)Isolation of endophytic bacteria from peaPea plants were collected from 4 collection points on the farm of Tibet Agriculture and Animal Husbandry College,and a total of 50 endophytic bacteria were purified by traditional isolation and culture methods.(2)Promoting potential of endophytic bacteria in peaThe effective phosphorus content,nitrogen content and secreted IAA content of the 50 isolated endophytic bacteria were determined by molybdenum-antimony anti-colorimetry,automatic Kjeldahl method and SalksowskiⅡ colorimetry.In the study of the growth promotion ability of these 50 endophytic bacteria,it was found that: 49 strains of organophosphorus-decomposing bacteria,the available phosphorus increment was 4.56～1086.25mg/L,and the strain with the largest available phosphorus increment was ML-5;48 strains of IAA bacteria,of which strain LT-1 has the strongest secretion capacity,with a secretion amount of 214.47mg/L;31 strains of nitrogen-fixing bacteria,of which strain MT-2 has the highest nitrogen fixation amount,reaching 3.60mg/L.Capable of dissolving phosphorus,fixing nitrogen and producing IAA.(3)Antagonistic effects of pea endophytic bacteriaAn antagonistic test was carried out on 50 strains of bacteria by flat-pane confrontation method.After preliminary screening and re-screening,11 strains were finally obtained,which had good resistance to wheat sheath blight,wheat root rot,wheat scab,corn leaf spot and tomato early blight The strain of action.Among them,strain Y-1 has a good antagonistic effect on these five pathogenic bacteria.The broad-spectrum inhibition rate of strain Y-1 against the five pathogenic bacteria was 63.4% at 24 h,71.6% at 48 h,and 75.8% at 96 h.(4)Growth promotion effect of pea endophytic bacteriaTen of the 50 endophytic bacteria isolated from Tibetan pea plants were selected for field experiments.The test results show that pea endophytic bacteria have different degrees and different aspects of promoting growth of pea plants.Compared with CK,the strain CT-2increased in plant height,root length,fresh weight,dry weight,nodule and chlorophyll at the initial flowering stage by 38.75%,24.36%,177.42%,44.07%,13.73%,and-4.35%.During the initial pod period,they increased by 134.78%,34.46%,196.54%,138.49%,50.61% and-6.97%,respectively.(5)Physicochemical characteristics of pea plantsThe contents of total nitrogen,total phosphorus and total potassium in pea plants at different growth stages were determined by the automatic Kjeldahl method,molybdenum-antimony colorimetry and atomic absorption spectrometry.Studies have shown that the content of total nitrogen,total phosphorus,and total potassium of CT-2 increased by30.02%,145.19%,and 71.22% compared with CK at the initial flowering stage,and increased by 5.64 compared to CK at the initial pod stage5.64%,-54.23% and 28.67%,the comprehensive evaluation ranking of membership function is the first.(6)Study on soil physical and chemical characteristicsAdopt automatic Kjeldahl nitrogen determination method,molybdenum antimony anti-colorimetry,atomic absorption spectrometry,alkali hydrolysis nitrogen distillation,molybdenum antimony anti-colorimetry,ammonium acetate-flame photometer method and p H meter Determination of total nitrogen,total phosphorus,total potassium,available nitrogen,available phosphorus,available potassium,organic matter,p H in soil.Studies have shown that the treatment with the highest content of organic matter and total potassium is CK1,which is an increase of 2.33% and 19.47% compared with CK;the treatment with the highest total phosphorus and available phosphorus content is CK;Compared with CK,it increased by 15.89%;the treatment with the highest available nitrogen and available potassium content was CT-2,which was increased by 69.06% and 164.11% compared with CK;the highest p H treatment was GM-1,and the lowest was LT-1.(7)Study on physiological and biochemical characteristics of strainsThe salt,acid,and alkali resistance of the strains were determined by setting the salt,acid,and alkali concentrations.The research showed that the optimal salt concentrations of the strains ML-3,L-3,and CT-2 were CK and 4%,respectively,CK,the optimum p H is p H7,p H7,p H5 respectively.The best carbon and nitrogen sources of the strains were screened by setting 5 different carbon sources and 5 different nitrogen sources.The research showed that the best carbon and nitrogen sources of strains ML-3 and L-3 were maltose and beef,respectively.The best carbon and nitrogen sources for the strain CT-2 were mannitol and peptone.By Gram staining,all 3 strains tested were Gram-positive.In the contact enzyme test,all three strains tested were positive.In the methyl red test,all three strains tested showed negative reactions.