基于根際營養(yǎng)利用能力的植物根際促生菌的篩選及促生效應(yīng)研究
發(fā)布時(shí)間：2018-08-09 18:01
【摘要】：優(yōu)良菌種是微生物肥料的基礎(chǔ),根際競爭能力是微生物肥料菌株發(fā)揮作用的前提,篩選的植物根際促生菌必須要能較好地適應(yīng)于植物的根際環(huán)境。本研究利用玉米、黃瓜、番茄三種作物的根系分泌物為初篩培養(yǎng)基的營養(yǎng)源,從相對應(yīng)作物的根際土壤樣品中分離篩選能利用根系分泌物快速生長的菌株,通過測定所篩菌株的促生特性,進(jìn)一步結(jié)合平皿幼苗促生試驗(yàn)復(fù)篩優(yōu)良菌株,并通過盆栽試驗(yàn)評價(jià)促生效果及對所篩菌株進(jìn)行系統(tǒng)鑒定。最后選擇JScG、SCmB、XJmG、SDtG四株芽孢桿菌進(jìn)行針對不同作物的根際定殖能力研究、液體發(fā)酵培養(yǎng)基組分優(yōu)化、根際促生微生物肥料研制并研究其盆栽促生效果。所獲主要結(jié)果如下:1.PGPR菌株初篩共獲得24株菌株,它們均可在以植物根系分泌物為唯一營養(yǎng)來源的培養(yǎng)基上迅速生長,且具有溶磷、產(chǎn)NH3、產(chǎn)IAA及產(chǎn)嗜鐵素等一種或多種促生性能。其中具有產(chǎn)氨能力的14株,占所篩菌株的58.3%;可產(chǎn)IAA的20株,占所篩菌株的83.3%,菌株JScB的IAA產(chǎn)量最大,達(dá)到了 73.28 mg L-1;可產(chǎn)鐵載體的12株,占所篩菌株的50%;具有溶解有機(jī)磷能力的15株,占所篩菌株的62.5%;所篩各菌株均有一定的溶解無機(jī)磷的能力,但差異很大,溶磷量范圍在19.14-200.05 mg L-1。根據(jù)平皿幼苗促生試驗(yàn),最終復(fù)篩出12株P(guān)GPR菌株。其中對玉米有促生作用的根際細(xì)菌為SCmB、SCmD、NMGmA、XJmG;對番茄有促生作用的根際細(xì)菌為SDtC、SDtG、GZtB、GZtE;對黃瓜有促生作用的根際細(xì)菌為HNcB、HNcH、JScB、JScG。2.將復(fù)篩得到的12株P(guān)GPR菌株用于盆栽試驗(yàn),分別回接至宿主植物玉米、黃瓜、番茄根際,生長40d后其株高、生物量、根系形態(tài)(總根長、根表面積等)和葉綠素含量等顯著高于不接菌對照,經(jīng)16SrDNA分子鑒定,5株為芽孢桿菌屬(Bacillus spp.);3株為劍菌屬(Ensifer spp.);2株為根瘤菌屬(Rhizobium spp.);1株為蒼白桿菌屬(Ochrobactrum sp.);1株為微桿菌屬(Microbacterium sp.)。其中,在玉米盆栽試驗(yàn)中,菌株SCmB促生效果最好,其次為XJmG;在番茄盆栽試驗(yàn)中,促生效果最好的菌株為GZtB,其次為SDtG;在黃瓜盆栽試驗(yàn)中,促生效果最好的菌株為HNcB,其次為JScB和JScG。上述促生效果好的7株菌中,有4株是芽孢桿菌,能產(chǎn)生耐熱抗逆的芽孢,是根際促生微生物肥料研發(fā)的理想菌種,故選擇SCmB、XJmG、SDtG、JScG四株芽孢桿菌進(jìn)行深入研究。3.測定了 JScG、SCmB、XJmG、SDtG四株芽孢桿菌的生物膜形成及對玉米、黃瓜、番茄三種作物的根際定殖能力。其中菌株SDtG和JScG的生物膜形成能力最強(qiáng),SCmB次之,XJmG最弱。在限菌砂培定殖試驗(yàn)中,SCmB菌株在玉米根際的定殖能力最強(qiáng),達(dá)到了 9.29×106CFUg-1根;SDtG和JScG在黃瓜根際的定殖能力最強(qiáng),分別達(dá)到了 4.69×106CFU g-1根和4.12×106CFU-1根;XJmG在番茄根際的定殖能力最強(qiáng),達(dá)到了 8.07× 1 06 CFU g-1 根。4.利用單因素試驗(yàn)和正交試驗(yàn)對四株芽孢桿菌的液體發(fā)酵培養(yǎng)基組分進(jìn)行了優(yōu)化,菌株JScG和SCmB液體發(fā)酵培養(yǎng)基的最優(yōu)配比為糖蜜3%,復(fù)合氮源0.6%,氯化鈉0.5%,氯化鈣0.1%,硫酸錳0.05%;XJmG最優(yōu)配比為糖蜜2.5%,復(fù)合氮源0.6%,氯化鈉0.5%,磷酸二氫鉀0.1%,氯化鈣0.1%,硫酸鎂0.1%,硫酸錳0.05%;SDtG最優(yōu)配比為糖蜜3%,復(fù)合氮源0.6%,氯化鈉0.25%。5.根據(jù)四株芽孢桿菌在液體發(fā)酵培養(yǎng)基中的生長曲線,選擇了 72 h的液體發(fā)酵液進(jìn)行了根際促生微生物肥的初步制作,即用腐熟堆肥吸附10%(v/w)液體發(fā)酵液制作四種微生物肥料,其芽孢數(shù)在5.0×108 CFU g-1干重,施用0.5%的微生物肥料于玉米和番茄根際,生長35 d后其株高、生物量、根系形態(tài)(總根長、根表面積等)等顯著高于對照,其中菌株SCmB對玉米的促生效果最好,菌株XJmG對番茄的促生效果最好。
[Abstract]:The excellent strain is the basis of microbial fertilizer, and the competitive ability of the rhizosphere is the prerequisite for the microbial fertilizer strain to play its role. The selected plant rhizosphere bacteria must be well adapted to the rhizosphere environment of the plant. This study uses the root secretions of three crops of maize, cucumber and tomato as the source of the primary culture medium. The strain of rapid growth of root exudates was isolated and screened in the rhizosphere soil samples. By measuring the growth promoting characteristics of the screened strains, the fine strains were screened by the seedling growth promoting test of Petri dish seedlings, and the effect of growth promotion and the system identification of the screened strains were evaluated by pot experiment. Finally, the four buds of JScG, SCmB, XJmG and SDtG were selected. The bacilli carried out the study on the rhizosphere colonization ability of different crops, the optimization of the medium composition of liquid fermentation medium, the development of rhizosphere microbial fertilizer and the effect of its pot cultivation. The main results were as follows: 24 strains of 1.PGPR strains were obtained at the beginning of screening, and they could be used in the culture medium with the sole nutrient source of plant root exudates. It is fast growing and has one or more growth promoting properties, such as phosphorus dissolving, producing NH3, producing IAA and ferric iron, which have 14 strains of ammonia production, accounting for 58.3% of the sieved strain, 20 of IAA and 83.3% of the screened strain, and the IAA yield of the strain JScB is 73.28 mg L-1, and 12 of the ferric carrier, 50% of the screened strain. 15 strains of organophosphorus, which accounted for 62.5% of the screened strains, all had a certain ability to dissolve inorganic phosphorus, but the difference was very large. The amount of phosphorus dissolved in 19.14-200.05 mg L-1. was finally screened out of 12 PGPR strains according to the growth promoting test of Petri dish seedlings. Among them, the Rhizosphere bacteria that have the promoting effect on maize were SCmB, SCmD, NMGmA, XJmG; and tomato. SDtC, SDtG, GZtB, GZtE, the rhizosphere bacteria that have the effect of promoting growth, and the rhizosphere bacteria that have a stimulating effect on cucumber are HNcB, HNcH, JScB, and JScG.2. will be used to resieve 12 strains of PGPR strains used for pot experiment, and the plant height, biomass, root morphology (total root length, root surface area, etc.) of the host plant maize, cucumber, tomato rhizosphere and growth 40d respectively. Chlorophyll content was significantly higher than that of non bacteria control. 5 strains were identified by 16SrDNA molecule (Bacillus spp.), 3 were rapiti (Ensifer spp.), 2 strains were rhizobia (Rhizobium spp.), 1 were pallid (Ochrobactrum sp.), and 1 strains were micro bacilli (Microbacterium sp.). In the pot experiment of maize, strain SCmB In the pot experiment of tomato, the best growth promoting effect was GZtB, followed by SDtG. In the pot experiment of cucumber, the best producing strain was HNcB, followed by 7 strains of JScB and JScG., the 7 strains were bacillus, which could produce the spores with heat resistance and resistance. Four strains of Bacillus, SCmB, XJmG, SDtG, and JScG were selected for further study on the ideal strain of fertilizer research and development. The biofilm formation of JScG, SCmB, XJmG, SDtG four strains of Bacillus and the rhizosphere colonization of three crops of maize, cucumber and tomato were determined by.3.. Among them, the biofilm of SDtG and JScG was the strongest, and the SCmB was the weakest. The colonization ability of SCmB strain in maize rhizosphere was the strongest, reaching 9.29 x 106CFUg-1 roots in maize rhizosphere; SDtG and JScG had the strongest colonization ability in Cucumber Rhizosphere, which reached 4.69 x 106CFU g-1 root and 4.12 x 106CFU-1 root respectively. XJmG had the strongest colonization ability in tomato rhizosphere, reaching 8.07 x 106 CFU g-1 root.4. utilization single. Factors test and orthogonal test were used to optimize the liquid fermentation medium of four Bacillus strains. The optimal ratio of strain JScG and SCmB liquid fermentation medium was molasses 3%, compound nitrogen source 0.6%, sodium chloride 0.5%, calcium chloride 0.1%, manganese sulphate 0.05%, XJmG optimum ratio was sugar honey 2.5%, compound nitrogen source 0.6%, sodium chloride 0.5%, potassium dihydrogen phosphate 0.1% Calcium chloride 0.1%, Magnesium Sulfate 0.1%, manganese sulphate 0.05%, SDtG optimum ratio of molasses 3%, compound nitrogen source 0.6%, sodium chloride 0.25%.5. based on the growth curve of four bacillus in liquid fermentation medium, selected 72 h liquid fermentation liquid to produce rhizosphere microbial fertilizer, that is to use decomposed composting for 10% (v/w) liquid fermentation. Four kinds of microbial fertilizers were made in the liquid, the number of spore was 5 x 108 CFU g-1 dry weight, and 0.5% microbial fertilizer was used in maize and tomato rhizosphere. After the growth of 35 d, the plant height, biomass, root morphology (total root length, root surface area and so on) were significantly higher than those of the control, among which strain SCmB had the best effect on maize growth, and the growth promoting effect of strain XJmG on Tomato Best.
【學(xué)位授予單位】：南京農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：S144;S182
[Abstract]:The excellent strain is the basis of microbial fertilizer, and the competitive ability of the rhizosphere is the prerequisite for the microbial fertilizer strain to play its role. The selected plant rhizosphere bacteria must be well adapted to the rhizosphere environment of the plant. This study uses the root secretions of three crops of maize, cucumber and tomato as the source of the primary culture medium. The strain of rapid growth of root exudates was isolated and screened in the rhizosphere soil samples. By measuring the growth promoting characteristics of the screened strains, the fine strains were screened by the seedling growth promoting test of Petri dish seedlings, and the effect of growth promotion and the system identification of the screened strains were evaluated by pot experiment. Finally, the four buds of JScG, SCmB, XJmG and SDtG were selected. The bacilli carried out the study on the rhizosphere colonization ability of different crops, the optimization of the medium composition of liquid fermentation medium, the development of rhizosphere microbial fertilizer and the effect of its pot cultivation. The main results were as follows: 24 strains of 1.PGPR strains were obtained at the beginning of screening, and they could be used in the culture medium with the sole nutrient source of plant root exudates. It is fast growing and has one or more growth promoting properties, such as phosphorus dissolving, producing NH3, producing IAA and ferric iron, which have 14 strains of ammonia production, accounting for 58.3% of the sieved strain, 20 of IAA and 83.3% of the screened strain, and the IAA yield of the strain JScB is 73.28 mg L-1, and 12 of the ferric carrier, 50% of the screened strain. 15 strains of organophosphorus, which accounted for 62.5% of the screened strains, all had a certain ability to dissolve inorganic phosphorus, but the difference was very large. The amount of phosphorus dissolved in 19.14-200.05 mg L-1. was finally screened out of 12 PGPR strains according to the growth promoting test of Petri dish seedlings. Among them, the Rhizosphere bacteria that have the promoting effect on maize were SCmB, SCmD, NMGmA, XJmG; and tomato. SDtC, SDtG, GZtB, GZtE, the rhizosphere bacteria that have the effect of promoting growth, and the rhizosphere bacteria that have a stimulating effect on cucumber are HNcB, HNcH, JScB, and JScG.2. will be used to resieve 12 strains of PGPR strains used for pot experiment, and the plant height, biomass, root morphology (total root length, root surface area, etc.) of the host plant maize, cucumber, tomato rhizosphere and growth 40d respectively. Chlorophyll content was significantly higher than that of non bacteria control. 5 strains were identified by 16SrDNA molecule (Bacillus spp.), 3 were rapiti (Ensifer spp.), 2 strains were rhizobia (Rhizobium spp.), 1 were pallid (Ochrobactrum sp.), and 1 strains were micro bacilli (Microbacterium sp.). In the pot experiment of maize, strain SCmB In the pot experiment of tomato, the best growth promoting effect was GZtB, followed by SDtG. In the pot experiment of cucumber, the best producing strain was HNcB, followed by 7 strains of JScB and JScG., the 7 strains were bacillus, which could produce the spores with heat resistance and resistance. Four strains of Bacillus, SCmB, XJmG, SDtG, and JScG were selected for further study on the ideal strain of fertilizer research and development. The biofilm formation of JScG, SCmB, XJmG, SDtG four strains of Bacillus and the rhizosphere colonization of three crops of maize, cucumber and tomato were determined by.3.. Among them, the biofilm of SDtG and JScG was the strongest, and the SCmB was the weakest. The colonization ability of SCmB strain in maize rhizosphere was the strongest, reaching 9.29 x 106CFUg-1 roots in maize rhizosphere; SDtG and JScG had the strongest colonization ability in Cucumber Rhizosphere, which reached 4.69 x 106CFU g-1 root and 4.12 x 106CFU-1 root respectively. XJmG had the strongest colonization ability in tomato rhizosphere, reaching 8.07 x 106 CFU g-1 root.4. utilization single. Factors test and orthogonal test were used to optimize the liquid fermentation medium of four Bacillus strains. The optimal ratio of strain JScG and SCmB liquid fermentation medium was molasses 3%, compound nitrogen source 0.6%, sodium chloride 0.5%, calcium chloride 0.1%, manganese sulphate 0.05%, XJmG optimum ratio was sugar honey 2.5%, compound nitrogen source 0.6%, sodium chloride 0.5%, potassium dihydrogen phosphate 0.1% Calcium chloride 0.1%, Magnesium Sulfate 0.1%, manganese sulphate 0.05%, SDtG optimum ratio of molasses 3%, compound nitrogen source 0.6%, sodium chloride 0.25%.5. based on the growth curve of four bacillus in liquid fermentation medium, selected 72 h liquid fermentation liquid to produce rhizosphere microbial fertilizer, that is to use decomposed composting for 10% (v/w) liquid fermentation. Four kinds of microbial fertilizers were made in the liquid, the number of spore was 5 x 108 CFU g-1 dry weight, and 0.5% microbial fertilizer was used in maize and tomato rhizosphere. After the growth of 35 d, the plant height, biomass, root morphology (total root length, root surface area and so on) were significantly higher than those of the control, among which strain SCmB had the best effect on maize growth, and the growth promoting effect of strain XJmG on Tomato Best.
【學(xué)位授予單位】：南京農(nóng)業(yè)大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：S144;S182
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