本文選題：叢枝菌根真菌　切入點(diǎn)：刺槐　出處：《西北農(nóng)林科技大學(xué)》2016年博士論文　論文類型：學(xué)位論文

【摘要】：本文通過黃土高原半干旱區(qū)5種主要林木刺槐(Robinia pseudoacacia)、杜松(Juniperus communis)、青楊(Populus cathayana)、沙棘(Hippophae rhamnoides)和旱柳(Salix matsudana)根際叢枝菌根真菌(Arbuscular mycorrhizal fungi,AMF)資源調(diào)查,分析了AMF與土壤因子的關(guān)系,研究了干旱脅迫下AMF對(duì)刺槐幼苗生長(zhǎng)、光合作用、抗氧化酶活性等生理生化指標(biāo)的影響,AMF對(duì)刺槐幼苗抗氧化酶基因和水孔蛋白基因表達(dá)的調(diào)控,初步揭示了AMF提高刺槐的抗旱機(jī)制。得出以下主要結(jié)果:1.黃土高原半干旱區(qū)AMF資源及其與土壤和氣候因子的相關(guān)性從刺槐、杜松、青楊、沙棘和旱柳5種林木根際土壤分離到8屬21種AMF:管柄囊霉屬(Funneliformis)7種,為優(yōu)勢(shì)屬,幼套近明囊霉(Claroideoglomus etunicatum)和網(wǎng)狀球囊霉(Glomus reticulatum)是優(yōu)勢(shì)種。土壤脲酶、過氧化氫酶和全磷是影響AMF總侵染率和孢子密度最直接的因子;轉(zhuǎn)化酶是影響AMF物種豐富度、Shannon-Wiener多樣性指數(shù)和Shannon均勻度指數(shù)最直接的因子。從6個(gè)不同半干旱區(qū)刺槐根際土壤分離得到9屬23種AMF:管柄囊霉屬6種,為優(yōu)勢(shì)屬,根內(nèi)根生囊霉(Rhizophagus intraradices)、兩型管柄囊霉(Fun.dimorphicum)、聚叢根生囊霉(R.aggregatum)、單孢管柄囊霉(Fun.monosporum)、凹坑管柄囊霉(Fun.multiforum)、地管柄囊霉(Fun.geosporum)和幼套近明囊霉是優(yōu)勢(shì)種。通徑分析表明降雨量是影響AMF狀況和多樣性指數(shù)最直接的因子。2.干旱脅迫條件下,AMF對(duì)刺槐光合作用的影響采用人工模擬干旱法研究根內(nèi)根生囊霉(R.irregularis)對(duì)刺槐生長(zhǎng)、葉綠素含量、氣體交換參數(shù)、葉綠素?zé)晒鈪?shù)等的影響。結(jié)果發(fā)現(xiàn),接種R.irregularis能夠提高刺槐葉綠素a、類胡蘿卜素、Fm、qP、NPQ和ΦPSII值,正常水分條件下,分別提高120.8%、44.7%、28.6%、3.4%、4.2%和7.0%;干旱脅迫條件下,分別提高69.5%、265.0%、12.8%、4.3%、3.8%和7.5%,與對(duì)照相比差異均達(dá)顯著水平。菌根化刺槐葉片凈光合速率和氣孔導(dǎo)度也顯著高于對(duì)照,正常水分條件下,分別增加16.4%和12.8%;干旱脅迫條件下,分別增加14.3%和8.1%。說明干旱脅迫條件下AMF與刺槐共生提高了宿主葉片PSII光化學(xué)活性與光合電子傳遞能力,保持高的光能利用效率與光合作用潛力,干旱脅迫條件下菌根化刺槐能更好地進(jìn)行光合作用。3.干旱脅迫條件下,AMF對(duì)刺槐抗氧化能力的影響采用人工模擬干旱法研究根內(nèi)根生囊霉(R.irregularis)對(duì)刺槐抗氧化能力的影響。結(jié)果發(fā)現(xiàn),接種R.irregularis后,刺槐根系O2、H_2O2和MDA含量降低,分別較對(duì)照顯著降低20.5%、33.1%和26.8%,說明接種AMF緩解了干旱脅迫對(duì)刺槐造成的氧化損傷。正常水分條件下,菌根化刺槐葉片和根系超氧化物歧化酶(SOD)、過氧化物酶(POD)、過氧化氫酶(CAT)、抗壞血酸過氧化物酶(APX)、谷胱甘肽還原酶(GR)活性分別增加53.6%和33.3%、20.4%和16.3%、20.9%和10.3%、67.4%和36.4%、2.7%和11.0%;干旱脅迫條件下,分別增加1.1%和27.2%、25.7%和23.5%、13.5%和9.8%、78.3%和58.2%、22.7%和14.4%,其中干旱脅迫條件下接種與對(duì)照葉片SOD、POD、CAT、APX和GR活性差異均達(dá)顯著水平。說明菌根化刺槐可以通過增強(qiáng)自身的抗氧化酶活性來清除干旱脅迫產(chǎn)生的活性氧。4.干旱脅迫條件下,AMF對(duì)刺槐抗氧化酶基因表達(dá)的影響采用實(shí)時(shí)熒光定量PCR(qRT-PCR)技術(shù)研究了R.irregularis對(duì)刺槐根、莖和葉片銅鋅超氧化物歧化酶基因(Cu/Zn-SOD)、抗壞血酸過氧化物酶基因(APX)和谷胱甘肽還原酶(GR)基因表達(dá)的影響。結(jié)果發(fā)現(xiàn),接種AMF上調(diào)了刺槐根、莖和葉片Cu/Zn-SOD、APX和GR基因的表達(dá)。正常土壤水分條件下,接種AMF上調(diào)了刺槐根、莖和葉片APX基因的表達(dá),分別為對(duì)照的3.31、3.17和3.06倍;接種AMF也上調(diào)了刺槐葉片GR基因的表達(dá),為對(duì)照的3.92倍。干旱脅迫條件下,接種AMF的刺槐根、莖和葉片Cu/Zn-SOD基因表達(dá)均表現(xiàn)為上調(diào),分別為對(duì)照的1.47、1.62和1.49倍;接種AMF也上調(diào)了刺槐根系GR基因的表達(dá),為對(duì)照的1.96倍。說明菌根化刺槐通過提高其根、莖和葉片Cu/Zn-SOD、APX和GR基因的表達(dá)來調(diào)控活性氧代謝,使其抗旱性增強(qiáng)。5.干旱脅迫條件下,AMF對(duì)刺槐水孔蛋白基因表達(dá)的影響采用qRT-PCR技術(shù)研究了接種R.irregularis對(duì)刺槐根、莖和葉片水孔蛋白(Aquaporin,AQP)基因表達(dá)的影響。從刺槐根部克隆得到RpTIP1;1、RpTIP1;3、RpTIP2;1、RpPIP1;1、RpPIP1;3和RpPIP2;1 6個(gè)水孔蛋白基因,6個(gè)RpAQPs cDNA全長(zhǎng)為825~1201 bp,含有750~870 bp開放閱讀框,編碼249~289個(gè)氨基酸,分子量為25.37~31.06 kDa,等電點(diǎn)為5.09~8.96,RpAQPs編碼的氨基酸序列與蒺藜苜蓿的同源性達(dá)89%~97%。組織表達(dá)分析表明,刺槐RpPIP基因在根部高表達(dá),而RpTIP基因(RpTIP1;3除外)在葉片高表達(dá)。干旱脅迫下,接種AMF的刺槐RpTIP2;1和RpPIP2;1基因在根中的表達(dá)表現(xiàn)出上調(diào)趨勢(shì),分別較對(duì)照提高54.7%和79.4%;在莖部,菌根化刺槐RpTIP1;1、RpTIP2;1和RpPIP2;1基因的相對(duì)表達(dá)量分別為對(duì)照的1.17、1.83和1.15倍;成熟葉片中,菌根化刺槐RpTIP2;1和RpPIP2;1基因相對(duì)表達(dá)量分別為對(duì)照的1.44和1.61倍。水孔蛋白家族基因在控制水分運(yùn)輸過程中發(fā)揮的作用不盡相同,接種AMF能通過促進(jìn)刺槐水孔蛋白基因的表達(dá)來增強(qiáng)植物對(duì)干旱脅迫的適應(yīng)性。
[Abstract]:The 5 main forest of Robinia pseudoacacia by semi arid area of Loess Plateau (Robinia pseudoacacia), Du Song (Juniperus communis), poplar (Populus cathayana), sea buckthorn (Hippophae rhamnoides) and willow (Salix matsudana) in rhizosphere of arbuscular mycorrhizal fungi (Arbuscular mycorrhizal, fungi, AMF) resource survey, analyzes the relationship between AMF and soil factors study on photosynthesis under drought stress, the growth of AMF, the effects of physiological and biochemical indexes of Robinia pseudoacacia seedlings, antioxidant enzyme activity, regulation of AMF on gene expression of antioxidant enzymes of Robinia pseudoacacia seedlings and aquaporin genes, revealed that the AMF mechanism to improve the drought resistance of Robinia pseudoacacia. The main results are as follows: correlation of AMF resources in semi arid region of the Loess Plateau and its 1. with soil and climate factors from locust, Du Song, poplar, willow and seabuckthorn 5 kinds of tree rhizosphere soil from 8 genera and 21 species of Trichoderma AMF: capsule tube handle (Funneliformis) for 7. The dominant genera, young set of near Ming (Claroideoglomus etunicatum) and fungus Glomus reticulate (Glomus reticulatum) is the dominant species. Soil urease, catalase and total phosphorus are the factors influencing AMF colonization and spore density directly; invertase was influenced by AMF factor Shannon-Wiener species richness, diversity index and Shannon evenness the index directly. 9 genera and 23 species of AMF: tube handle pouch mildew genera and 6 species isolated from the rhizosphere soil of Robinia pseudoacacia in 6 different semi arid region, was the dominant genus in the roots, the root fungus (Rhizophagus intraradices), type two (Fun.dimorphicum) tube handle pouch mildew, leaf clumping fungus (R.aggregatum) the tube handle, single spore fungus (Fun.monosporum), pit tube handle pouch mildew (Fun.multiforum), pipe handle pouch mildew (Fun.geosporum) and the set of near Ming capsule mold is dominant. The path analysis showed that rainfall is the impact of AMF status and diversity index of the most direct factor.2. stem Under drought stress, artificial drought of root root fungus with AMF on Photosynthesis of Robinia pseudoacacia (R.irregularis) on the growth of Robinia pseudoacacia, chlorophyll content, gas exchange parameters, the effect of chlorophyll fluorescence parameters. The results showed that R.irregularis inoculation can improve the locust chlorophyll a, carotenoid, Fm, qP, NPQ and with PSII, under normal water condition, increased by 120.8%, respectively, 44.7%, 28.6%, 3.4%, 4.2% and 7%; under drought stress, increased by 69.5%, respectively, 265%, 12.8%, 4.3%, 3.8% and 7.5%, compared with the control difference reached significant level. The mycorrhizal locust leaf net photosynthetic rate and stomatal conductance was significantly higher than that in control, under normal water condition, an increase of 16.4% and 12.8% respectively; under drought stress, respectively increased 14.3% and 8.1%. AMF and Robinia pseudoacacia under drought stress increased leaf PSII symbiotic host photochemical activity and photosynthetic power Sub transmission capacity, maintain the high light energy utilization efficiency and photosynthetic potential of mycorrhizal fungi of Robinia pseudoacacia under drought stress to.3. photosynthesis under drought stress, artificial drought of root root fungus with AMF on antioxidant capacity of black locust (R.irregularis) effects on antioxidant capacity of Robinia pseudoacacia. The results showed that R.irregularis after inoculation, Robinia pseudoacacia roots decreased H_2O2 and O2, MDA content, were significantly lower than the CK 20.5%, 33.1% and 26.8%, AMF inoculation alleviated the drought stress on oxidative damage caused by normal locust. Water conditions, mycorrhizal locust leaves and roots of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), glutathione reductase (GR) activity were increased by 53.6% and 33.3%, 20.4% and 16.3%, 20.9% and 10.3%, 67.4% and 36.4%, 2.7% and 11% ; under drought stress condition, increased by 1.1% and 27.2% respectively, 25.7% and 23.5%, 13.5% and 9.8%, 78.3% and 58.2%, 22.7% and 14.4%, which under drought stress inoculation and control leaves SOD, POD, CAT, APX and GR activity significantly. The mycorrhizal Robinia pseudoacacia through its antioxidant activity to remove the drought stress by reactive oxygen.4. drought stress condition, the effect of AMF on locust antioxidant enzyme gene expression by real-time fluorescent quantitative PCR (qRT-PCR) of R.irregularis on locust roots, stems and leaves of copper zinc superoxide dismutase gene (Cu/Zn-SOD), ascorbate peroxidase gene (APX) and glutathione reductase (GR) gene expression in vitro. The results showed that AMF inoculation increased locust root, stems and leaves of Cu/Zn-SOD, the expression of APX and GR genes. The normal soil moisture conditions, inoculation of AMF raised locust roots, stems and The expression of APX gene in leaves, respectively 3.31,3.17 and 3.06 times of inoculation; AMF also upregulated the expression of GR gene of Robinia pseudoacacia leaves, was 3.92 times of the control. Under drought stress, Robinia pseudoacacia roots inoculated with AMF, the expression of stem and leaf of Cu/Zn-SOD gene were up-regulated, respectively, according to 1.47,1.62 and 1.49 times AMF; inoculation increased the expression of GR gene of Robinia pseudoacacia roots, was 1.96 times of the control. The mycorrhizal Robinia pseudoacacia by improving its roots, stems and leaves of Cu/Zn-SOD, the expression of APX and GR genes to regulate the metabolism of active oxygen, which enhanced.5. drought resistance under drought stress, effect of AMF on the expression of aquaporin gene of Robinia pseudoacacia the study of locust root inoculation of R.irregularis using qRT-PCR technology, stems and leaves of aquaporins (Aquaporin, AQP) gene expression of RpTIP1 from locust Root Clone; 1, RpTIP1; 3, RpTIP2; 1, RpPIP1; 1, RpPIP1; 3 and RpPIP2; 16 Aquaporin 6, RpAQPs cDNA was 825~1201 BP, containing a 750~870 open reading frame of BP encoding 249~289 amino acids, molecular weight of 25.37~31.06 kDa, the isoelectric point was 5.09~8.96, RpAQPs encoding the amino acid sequence of Medicago truncatula homology of 89%~97%. tissue expression analysis showed that RpPIP gene is highly expressed in the roots of black locust, and RpTIP gene (RpTIP1; except 3) is highly expressed in leaves. Under drought stress, Robinia pseudoacacia inoculated with AMF RpTIP2; 1 and RpPIP2; 1 gene expression in root showed upward trend, respectively increased 54.7% and 79.4%; in the stems, mycorrhizal locust RpTIP1; 1, RpTIP2; 1 and RpPIP2; the relative expression of 1 genes were controlled 1.17,1.83 and 1.15 times; in mature leaves of mycorrhizal locust RpTIP2; 1 and RpPIP2; 1 gene expression levels were decreased to 1.44 and 1.61 times. The aquaporin family genes in the control of water transport process The role of AMF is different, and inoculation can enhance the adaptability of plants to drought stress by promoting the expression of the Robinia Robinia water pore protein gene.

【學(xué)位授予單位】：西北農(nóng)林科技大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：S792.27
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本文編號(hào)：1597211