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1.
香蕉果实特异性ACC合成酶基因的克隆及反义载体的构建   总被引:4,自引:0,他引:4  
乙烯是一种结构简单、功能多样的植物激素,它在植物的整个生长发育(包括种子萌发、花开花谢、果实成熟、衰老、器官脱落及胁迫反应等)过程中都起调控作用[1,2].ACC合成酶(EC.4.4.14)是植物体内乙烯生物合成的限速酶,对跃变型果实而言,当ACC合成酶基因的表达受抑制时,乙烯的生成量下降,果实成熟受阻,使贮藏期得以延长[3].现已从许多植物中克隆到了ACC合成酶基因[4].香蕉(MusaAAA Cavendish Subgroup)是典型的跃变果实,从香蕉中克隆ACC合成酶基因的目的,是利用其反义基因转化香蕉以培育耐贮藏的香蕉新品种并进行香蕉采后的分子生物学研究.  相似文献   

2.
卡特兰ACO基因克隆与反义表达载体的构建   总被引:1,自引:1,他引:0  
以卡特兰(Cattleya)花瓣为试材,提取其总RNA,并根据其他兰花的ACC氧化酶(1-aminocyclopropane-1-carboxylic acid oxidase, ACO)基因保守序列设计一对特异性引物,通过RT-PCR法克隆得到1条967bp的卡特兰ACO cDNA片断,共编码321个氨基酸残基。序列分析结果显示该克隆片断与已发表的其他兰花的ACO基因序列同源性很高,均在85%以上,尤其与原生种和其近亲属的同源性在95%以上。将克隆的卡特兰ACO片段反向连接到植物表达载体pBI121中CaMV35S启动子的下游,构建了卡特兰ACO基因的反义表达载体pBI121ACC,为进一步应用反义技术培养长花期卡特兰新品种奠定了基础,也为首次应用生物技术延长卡特兰花期做出了尝试。  相似文献   

3.
猕猴桃ACC合成酶基因家族四个成员的克隆   总被引:5,自引:0,他引:5  
通过PCR方法从中华猕猴桃中分离出ACC合成酶基因家族的四个成员(AC-ACS1A、AC-ACS1B、AC-ACS2和AC-ACS3)的基因组DNA片段,AC-ACS1A、AC-ACS1B和AC-ACS2与其它植物该基因的氨基酸序列同源性最高可达76%以上,而AC-ACS3与其它植物ACC合成酶基因的氨基酸序列同源性均低于60%,与已知的其它猕猴桃ACC合成酶基因的同源性在51%-56%之间,且不存在MSSFGL保守区,因而属于一个未见报道的新成员。  相似文献   

4.
据已报道的香石竹氨基环丙烷羧酸(ACC)氧化酶基因的cDNA序列设计特异引物,以香石竹品种“Master”基因组DNA为模板,用PCR方法克隆出香石竹ACO基因的部分序列。测序结果显示,克隆的序列与已报道序列基本一致。将克隆的香石竹ACO片段反向连接到植物表达载体pCAMBIA 1301中CaMV 35S启动子的下游,构建了香石竹ACO基因的反义表达载体pCAM-ACO2,为进一步通过反义技术延长转基因香石竹的花期奠定了基础。  相似文献   

5.
ACC对不同氮效率油菜生长后期硝态氮再利用的调控机理   总被引:1,自引:1,他引:0  
【目的】研究了进一步解析乙烯对油菜生长后期硝态氮 (NO3–) 再利用的影响,揭示植株生长后期氮素再利用的生理机制。【方法】以氮高效油菜品种湘油15 (27号) 与氮低效油菜品种814 (6号) 为试验材料,在15 mmol/L氮水平下,每7天浇灌一次50 mL 100 μmol/L 1-氨基环丙烷-1-羧酸 (1-am-inocyclopropane-1-carboxylic acid,简称ACC),研究ACC对植物生长后期 (花期、收获期) 氮素再利用的影响及其与氮素利用效率 (NUE) 的关系。并用拟南芥野生型 (col.0) 和突变体 (nrt1.5) 材料作为验证,分别于玻璃顶网室和22℃恒温培养室进行砂培试验。【结果】ACC处理显著抑制了油菜BnNRT1.5的表达,且植株的衰老可以显著诱导BnNRT1.5的表达。相对于对照处理,ACC处理植株韧皮部汁液NO3– 的再转运能力显著降低,导致下部叶NO3– 含量显著升高,中部叶NO3– 含量显著下降,上部叶NO3– 含量无显著变化,进而导致植株含氮量和籽粒含氮量显著提高,以及以生物量和籽粒产量为基础的氮素利用效率 (NUE) 显著降低。由此推测,油菜生长后期氮素的再利用能力受到NRT1.5基因的显著调控。拟南芥野生型和突变体材料的验证结果表明,相对于拟南芥野生型 (col.0) 材料,拟南芥nrt1.5植株生长后期相对于col.0有更多的NO3– 累积在植株衰老叶片中,更少的NO3– 通过韧皮部转运到生长旺盛的新叶,植物生长后期氮素从老叶向新叶转运的再利用能力显著降低。【结论】油菜生长后期氮素的再利用能力受到ACC的显著调控,油菜和拟南芥NRT1.5基因表达量分别受到抑制或者发生基因突变时,会导致植株韧皮部汁液NO3– 再转运量减少,更多NO3– 累积在衰老叶片中而不能得以高效的再利用。因此,调控油菜生长后期NRT1.5的表达,提高油菜生长后期氮素的再转运和利用可以作为提高氮素利用效率的有效手段。  相似文献   

6.
李明源  王继莲  田世梅 《核农学报》2023,(11):2151-2157
为探究盐碱区生态环境改良方式,丰富产1-氨基环丙烷-1-羧酸(ACC)脱氨酶菌种资源,从新疆克孜勒苏柯尔克孜自治州盐碱地盐爪爪(Kalidium foliatum)根际土壤中筛选产ACC脱氨酶菌株,结合16S rRNA基因测序对优良菌株鉴定,通过盆栽接种试验验证其促生效果。结果从盐爪爪根际分离到26株产ACC脱氨酶菌株,酶活性为0.4~5.6 U·mg-1,以PM14、PM16和PM24活性最高。经鉴定,菌株PM14、PM16和PM24分别归属肠杆菌属(Enterobacter)、赖氨酸芽孢杆菌属(Lysinibacillus)和假单胞菌属(Pseudomonas)。盆栽试验表明,接种3株菌对非盐碱胁迫下拟南芥地上(株高、地上干重)和地下(根长、根干重)部分生长均有明显促进作用;对盐碱土栽培小麦的地上部分(株高、茎粗、鲜重和干重、叶绿素含量)和根干重也有积极影响,尤以PM16最显著。PM14和PM16兼具解磷、固氮和分泌吲哚-3-乙酸(indole-3-acetic acid, IAA)的能力,PM24兼具固氮和分泌IAA的能力,均具有多种促生功能。本研究结果为盐...  相似文献   

7.
为筛选能产生1-氨基环丙烷-1-羧基(ACC)脱氨酶的大豆植物根际细菌,从大豆根际土壤中分离得到10株大豆根际细菌,其中5株在DF和ADF培养基中能以ACC作为唯一氮源生长.通过16S rDNA测序及生理生化实验证实,H1为地衣芽孢杆菌(Bacillus licheniformis),H2为苏云金杆菌(Bacillus...  相似文献   

8.
以1-氨基环丙烷-1-羧酸(ACC)为唯一氮源,从石油污染的土壤中分离得到一株具有ACC脱氨酶活性的菌株D5A.该菌在以ACC为唯一氮源条件下,其ACC脱氨酶比活力为0.084 U/mg.另外,D5A还具有产生吲哚乙酸(IAA)、耐盐以及溶磷的特性.在液体培养条件下该菌株产IAA高达112 mg/L,在90 g/kg盐含量的培养基中仍能够正常生长且具有较强的溶解矿物磷能力.同时该菌对酸碱具有良好的适应性,在初始pH4~10的LB培养基中生长良好.种子发芽试验表明在3 g/kg和6 g/kg浓度NaC1的逆境条件下,该菌株能显著提高高羊茅种子的发芽率和芽长.最后,通过对其进行生理生化特性和16S rDNA序列分析,该菌株初步鉴定为克雷伯氏菌(Klebsiellasp.).  相似文献   

9.
从新疆五家渠103团盐碱地盐穗木根际的3份土样中筛选出10株产ACC脱氨酶的菌株,通过对这10株菌株的ACC脱氨酶活性和耐盐能力的测定,结果显示其中有4株菌株ACC脱氨酶活性较高,且能在盐(NaCl)浓度12%生长。根据ACC脱氨酶活性以及耐盐性能的测定结果,选择W5、W8、W9这3株菌株进行鉴定,通过生理生化实验、Biolog鉴定、16S rDNA测序分析后,鉴定结果显示W5、W8为乳酪短杆菌(Brevibacterium casei),W9为普沙根瘤菌(Rhizobium pusense)。  相似文献   

10.
从盐生植物根际土中分离得到4株含1-氨基环丙烷-1-羧酸(ACC)脱氨酶的植物促生菌(PGPR),通过无菌育种袋栽培试验,考查其在不同盐分条件下对燕麦和黑麦草幼苗的促生效应。结果表明,4株菌对5 g/kg或10 g/kg NaCl盐分胁迫下的燕麦和黑麦草幼苗均表现出显著地缓解促生效应,其中假单胞菌属S1最显著,10 g/kg NaCl比无NaCl时促生作用更大。4株PGPR的ACC脱氨酶活性与植物生长参数(根长和下胚轴长)之间具有极显著的正相关性(Pearson相关系数>0.81)。  相似文献   

11.
Plants and bacteria can interact with one another in a variety of different ways. The interaction may be beneficial, harmful or neutral for the plant, and sometimes the impact of a bacterium may vary as the soil conditions change. While a number of different soil bacteria are phytopathogenic, the majority of the more agronomically important plant disease-causing soil microorganisms are fungi. On the other hand, plant growth-promoting bacteria are typically of three general types: those that form a symbiotic relationship with the plant, those that are endophytic and colonize the inner tissues of the plant, and those of soil bacteria, which have competitive abilities to colonize efficiently the rhizosphere and the surface of plant roots.While there have been significant advances in elucidating the mechanistic details of plant-bacterial interactions in recent years, many fundamental questions about these processes remain. Unfortunately, studies that focus on only a single biochemical pathway or mechanism often miss the multiplicity of effects that plants and bacteria have on one another, motivating the employment of broader proteome-wide approaches. On the other hand, using proteomics technology including high-resolution two-dimensional gel electrophoresis (2-DE) and high-sensitivity mass spectrometry (MS), it is possible to gain greater insight into the detailed impact that plants and soil bacteria have on one another.In this regard, of all of the proteomic studies of plant-bacterial interactions, the symbiotic interaction between nitrogen-fixing bacteria and legumes has been studied in the greatest detail. Studies of the proteome of plant-pathogen interactions have also received considerable attention. However, there are currently very few proteomic studies of endophytic and rhizosphere associated plant growth promoting bacteria.Here, some fundamental proteomic tools are introduced and the applications of one of these approaches (i.e., 2-DE coupled to MS) to the study of plant-bacterial interactions are discussed. This review specifically addresses the questions: what are the impacts of plants on the bacterial proteomes, and vice versa?  相似文献   

12.
从阴沟肠杆菌(Enterobactercloacae)UW4菌株提取细菌总DNA,根据已报道的阴沟肠杆菌ACC脱氨酶(1-aminocy-clopropane-1-carboxylicaciddeaminase)基因序列设计简并引物,通过PCR扩增获得1条约1.02kb特异片段,把该片段连接到pGEM-Tvector上进行测序。序列分析结果表明,该基因编码区全长为1017bp,共编码338个氨基酸残基,与报道的序列相比仅存在8个核苷酸的差异,同源性达99.1%;将其翻译成氨基酸序列后发现,仅引起了4个氨基酸残基的变化,氨基酸同源性为98.2%。利用DNASIS软件对蛋白质二级结构进行分析比较,发现其蛋白质二级结构及其单元与报道的序列完全一致。将其插入原核表达载体pET-28b进行诱导表达,发现其表达蛋白在分子量约39kD处比对照多出一明显条带;经Westernblot分析检测,发现此带即为His-ACDS融合蛋白。  相似文献   

13.
14.
The use of plant growth-promoting rhizobacteria (PGPR) as agricultural inputs for increasing crop production needs the selection of efficient bacteria with plant growth-promoting (PGP) attributes. Therefore, the purpose of this study was to evaluate the effects of 20 multi-traits bacteria on tea growth, nutrient uptake, chlorophyll contents, and enzyme activities under field conditions for over 3 years. These isolates were screened in vitro for their PGP traits such as the production of indole acetic acid (IAA), nitrogenase activity, phosphorus (P) solubilization, and 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity. Screening of rhizobacteria that show multiple PGP traits suggests that they stimulated overall plant growth, including shoot development and leaf yield, improving macro- and micro-nutrient uptake, chlorophyll contents, and activities of enzymes of tea plant. Use of strains with multiple PGP traits could be a more effective approach and have great potential for the environmentally-friendly tea production.  相似文献   

15.
The effects of the application of the jasmonic acid derivative n-propyl dihydrojasmonate (PDJ) on ethylene biosynthesis, volatile compounds, and endogenous jasmonic acid (JA) and methyl jasmonate (MeJA) were examined in Japanese apricot (Prunus mume Sieb.) infected by a pathogen (Colletotrichum gloeosporioides). The fruit were dipped into 0.4 mM PDJ solution before inoculation with the pathogen and stored at 25 °C for 6 days. The inoculation induced an increase in 1-aminocyclopropane-1-carboxylic acid (ACC), ethylene, JA, and MeJA. In contrast, PDJ application reduced the endogenous JA, MeJA, and ethylene production and expression of the ACC oxidase gene (PmACO1) caused by the pathogen infection. The lesion diameter with C. gloeosporioides decreased upon PDJ application. The alcohol, ester, ketone, and lactone concentrations and alcohol acyltransferase (AAT) activity increased in the pathogen-infected fruit, but were decreased by PDJ application. These results suggest that PDJ application might influence ethylene production through PmACO1 and that aroma volatile emissions affected by pathogen infection can be correlated with the ethylene production, which is mediated by the levels of jasmonates.  相似文献   

16.
丙二烯氧化物合酶(allene oxide synthase,AOS)是茉莉酸合成途径中第一个特异性的酶。本研究从"嘎啦"苹果中克隆了一个AOS的基因序列,将其命名为MdAOS,并对其进行了结构和表达分析。MdAOS基因编码区共有1 605 bp碱基,编码531个氨基酸,等电点为4.92,分子量为134.9 kD。MdAOS属于细胞色素P450基因家族的CYP74A亚类,并具有典型的P450成员特征,其编码的蛋白包含4个保守区域,分别为:Heme-Binding结构域、Ⅰ-Helix区、ETLR基序和PEEF基序。MdAOS蛋白的N末端有典型叶绿体信号肽。同源性分析表明MdAOS与碧桃(Amygdalus persica var.persica f.duplex.)AOS基因的同源性很高,且聚类结果也与双子叶植物聚为一类。实时荧光定量PCR分析结果显示,MdAOS在苹果各组织中均有表达且在茎中表达量最高。MdAOS能响应茉莉酸、水杨酸、脱落酸和乙烯的诱导,并在机械伤处理时表达上调。  相似文献   

17.
One novel banana fruit ripening related 1-aminocyclopropane-1-carboxylate (ACC) oxidase gene quite different from ACC oxidase genes from other species was cloned. In contrast to other studies, the polypeptide encoded by this gene, named Mh-ACO1, lacks the putative leucine zipper motif which is conserved in all known ACC oxidases including the other previously reported banana ACC oxidase, Mh-ACO2. The locus consists of two nearly identical paralogous ACC oxidase genes arranged in opposite orientation and separated by a 3.1-kb intergenic region. The has only two introns, at positions identical to , which comprises a coding region interrupted by three introns. The predicted amino acid sequence of Mh-ACO1 shares less than 50% identity to those of ACC oxidase from other climacteric fruits, while that of Mh-ACO2 shows more than 65% homology. When expressed in Saccharomyces cerevisiae -encoded protein possessed the enzyme activity for ethylene conversion. The levels of mRNA corresponding to both and increased during fruit ripening and were induced by exogenous ethylene. We conclude that both and contribute to increased ethylene production in fruits and these two genes are differentially expressed in fruits and other organs in banana.  相似文献   

18.
Application of plant growth-promoting bacteria(PGPB)is an environmentally sustainable option to reduce the effects of abiotic and biotic stresses on plant growth and productivity.Three 1-aminocyclopropane-1-carboxylic acid(ACC)deaminase-producing drought-tolerant bacteria were isolated from a rain-fed agriculture field in the Central Himalaya of Kumaun region,Uttarakhand,India and evaluated for their efficiency in improving finger millet(Eleusine coracana(L.)Gaertn.)plant growth under non-stressed and drought-stressed conditions.These bacteria withstood a substrate metric potential of -1.0 MPa(30% polyethylene glycol 8000)and therefore were considered drought-tolerant.These strains were identified as Pseudomonas spp.by fatty acid methyl ester analysis and 16S rRNA gene sequencing.The ACC deaminase activity of these strains was characterized at the biochemical level,and the presence of acd S gene,the structural gene for ACC deaminase,was confirmed by the polymerase chain reaction.Two sets of pot trials in glass house were set up,one for normal(non-stressed)and the other for drought-stressed conditions.After 5 weeks,one set of plants was subjected to drought stress for 5 d,while the other set continued to be watered.The same growth parameters were recorded for both sets of plants after 40 d of plant growth.The results of pot trials showed that treatments inoculated with ACC deaminase-producing bacterial strains significantly improved the growth performance of finger millet plants and foliar nutrient content as compared to uninoculated treatments under both non-stressed and drought-stressed conditions.In addition,a significant increase in antioxidant activity was observed,wherein bacterial stain inoculation improved plant fitness by protecting it from oxidative damage induced by drought.  相似文献   

19.
Wounding is one of the most effective stress signals to induce ethylene synthesis in persimmon (Diospyros kaki Thunb.). We found that wound-induced ethylene biosynthesis is subjected to negative feedback regulation in mature 'Saijo' persimmon fruit since ethylene production was enhanced by 1-methylcyclopropene (1-MCP) (an inhibitor of ethylene perception) pretreatment, which was approximately 1.8 fold of that in control tissues (without 1-MCP pretreatment). Wound-induced 1-aminocyclopropane-1-carboxylate (ACC) synthase activity and DK-ACS2 gene expression were substantially increased by 1-MCP pretreatment after 12 h, which resulted in much higher ACC content in 1-MCP pretreated tissues than that in a control after 24 h. These results indicated that wound-induced DK-ACS2 gene expression was negatively regulated by ethylene in mature persimmon fruit. However, 1-MCP pretreatment had no effect on DK-ACO1 gene expression, suggesting the independence of wound-induced DK-ACO1 on ethylene. Out of accord with DK-ACO1 gene expression, ACC oxidase activity was enhanced 48 h after wounding in 1-MCP pretreated tissues, reaching a peak 1.5-fold higher than that in control tissues at 60 h.  相似文献   

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