Genetic regulation of conidiation in Trichoderma hamatun

Achieving a balance between vegetative growth and spore production is essential for successful biocontrol by fungi. Low sporulation rates in the field can result in poor establishment and survival,whereas failure of conidia to recognise hosts can lead to persistence without efficacy. Commercial biocontrol products involve bulk preparations of conidia, however considerable variability in conidiation rates exists between biocontrol agents, which can restrict choice of strain for production. The majority of studies on Trichoderma conidiation have focused on the species T. viride and T. atroviride.These species form conidia in response to blue and near-UV light and/or nutrient deprivation and conidiation proceeds in a highly co-ordinated fashion, however relatively little is known on the genetic basis of Trichoderrma conidiation. In addition, whilst photoconidiation appears to be a general response detailed studies in other Trichoderma species are absent. In this study, conidiation in the lesser known biocontrol species T. hamatum is being investigated using a combined morphological and molecular approach. In contrast to T. atroviride, conidiation in response to blue-light was weaker and variable and suggested that additional triggers may be required for the T. hamatum photoresponse. A series of comparative photoconidiation assays are currently being undertaken investigating the effect of inoculum type and abiotic factors on timing and intensity of the response.Results will be discussed in relation to the current knowledge on conidial morphogenesis in Trichoderma. In addition to these morphological assays, a selection of genes implicated in sporulation and the blue-light responses are currently being isolated and characterised from T. hamatum. Two genes, phr1 and cmp1 , which were isolated previously from T. atroviride will be used as early and late markers of gene expression during the photoresponse in T. hamatum in order to define time points for harvesting comparable stage-specific RNA from T. hamatum and T. atroviride. Using degenerate PCR putative sporulation gene orthologues have also been identified in T. hamatum.Work is currently underway to isolate genomic clones of these genes from T. hamatum and T.atroviride. Sequence and expression analysis of orthologues, including expression in response to abiotic factors will be presented and discussed in relation to the current knowledge of the molecular basis of conidiation in Trichoderma and other filamentous fungi.……     

华山松疱锈病的重寄生真菌(深绿木霉)中几丁质酶基因cDNA片段的克隆

[目的]分离深绿木霉S5003中的几丁质酶基因,为获得高抗病的转基因植株奠定基础。[方法]采用华山松疱锈病的锈孢子壁作为诱导物,诱导深绿木霉SS003中几丁质酶基因的表达,并通过lit—PER方法扩增得到几丁质酶基因片段。[结果]锈孢子壁可诱导出高活性（40．17μg／10min）的几丁质酶,PCR扩增得到了长度为834bp的特异片段,经序列测定及分析显示该片段为几丁质酶基因片段。[结论]深绿木霉SS003的几丁质酶基因片段的获得,为分离全长基因以及进一步利用该基因生产几丁质酶以及进行基因转化提供了可能。     

华山松疱锈病的重寄生真菌(深绿木霉)中几丁质酶基因cDNA片段的克隆(英文)

[Objective] The aim of this study was to isolate chitinase gene from Trichoderma atroviride strain SS003. [Method] With the aeciospore wall of armandii pine blister rust as inducer, chitinase gene was induced to express in Trichoderma atroviride cells. The cDNA fragment of chitinase gene was cloned by RT-PCR approach. [Result] The activity of chitinase induced reached 40.17 μg/10 min; and the specific fragment amplified was 834 bp in length and proved to be the fragment of chitinase gene by sequencing and sequence analysis. [Conclusion] The result showed the feasibility of isolating the full length of chitinase gene and its transformation, and further producing chitinase.     

Study of signal transduction factors involved in mycoparasitic response of Trichoderma atroviride

Numerous Trichoderma spp. are mycoparasites and commercially applied as biological control agents against a large number of plant pathogenic fungi. The mycoparasitic interaction is host-specific and several research strategies have been applied to identify the main genes and compounds involved in the antagonist-plant-pathogen three-way interaction. During mycoparasitism, signals from the host fungus are recognised by Trichoderma, stimulating antifungal activities that are accompanied by mo…     

Supplemental blue and red light promote lycopene synthesis in tomato fruits

Lycopene, one of the strongest natural antioxidants known and the main carotene in ripe tomato, is very important for human health. Light is well known to be one of the most important environmental stimuli influencing lycopene biosynthesis; specifically, red light induces higher lycopene content in tomato. However, whether blue light promotes lycopene synthesis remains elusive and exactly how light stimulation promotes lycopene synthesis remains unclear. We applied supplemental blue and red lighting on tomato plants at anthesis to monitor the effect of supplemental blue and red lighting on lycopene synthesis. Our results showed that supplemental blue/red lighting induced higher lycopene content in tomato fruits; furthermore, we found that the expression of key genes in the lycopene synthesis pathway was induced by supplemented blue/red light. The expression of light signaling components, such as red-light receptor phytochromes (PHYs), blue-light receptor cryptochromes (CRYs) and light interaction factors, phytochrome-interacting factors (PIFs) and ELONGATED HYPOCOTYL 5 (HY5) were up- or down-regulated by blue/red lighting. Thus, blue and red light increased lycopene content in tomatoes by inducing light receptors that modulate HY5 and PIFs activation to mediate phytoene synthase 1 (PSY1) gene expression. These results provide a sound theoretical basis for further elucidation of the light regulating mechanism of lycopene synthesis in tomatoes, and for instituting a new generation of technological innovations for the enhancement of lycopene accumulation in crop production.     

Variations on conserved signaling pathways in biocontrol and development:G protein and MAPK genes of Trichoderma. atroviride and T. virens

Filamentous fungi employ conserved eukaryotic signaling pathway to detect and respond to environmental signals, including the presence of the host. Genetic experiment in which a particular signaling protein is lost, or its activity enhanced, have defined some of the function of heterotrimeric G proteins and MAP kinases in development and virulence. A hallmark of these studies is that orthologs in different species may have different functions. Antagonistic fungal-fungal interactions form …     

Cloning of first abc transporter encoding gene from Trichoderma spp. and its expression during stress and mycoparasitism

Trichoderma in its natural environment competes for nutrient uptake and is required to protect itself from adverse natural toxic compounds, such as those produced by plants and other microbes in the soil community, or synthetic toxic compounds released human activity. One of the most important metabolic pathways for drug resistance and substrate uptake, both in prokaryotes and eukaryotes, is ATP dependent. The role of ABC transporter proteins in the biology of Trichoderma is still not kno…     

深绿木霉(Trichoderma atroviride)T23降解有机磷农药敌敌畏转运蛋白TaPdr2基因的克隆与功能预测分析

根据前期研究,进行了对比分析后明确了深绿木霉中7个敌敌畏耐受相关ABC转运蛋白,其中TaPdr2基因是在短时间内对敌敌畏胁迫应答最为明显的一个基因。本研究通过简并引物同源克隆,在深绿木霉T23中克隆出TaPdr2基因,并对其全长序列进行了测序;明确了其外显子序列和内含子序列;构建了TaPdr2的系统发生树,发现深绿木霉TaPdr2与木霉属其他种PDR5亚家族ABC转运蛋白有很高的同源性,通过生物信息学分析后预测TaPdr2蛋白的ATPase及部分跨膜结构域序列相对保守且具有11个跨膜结构域;通过构建TaPdr2的敲除载体pC1300qh-F,为后续研究TaPdr2蛋白对麦角甾醇和鞘脂类物质含量影响的研究奠定了基础。     

Genetic improvement of Trichoderma ability to induce systemic resistance

The beneficial applications of Trichoderma spp. in agriculture include not only the control of plant pathogens, but also the improvement of plant growth, micronutrient availability, and plant tolerance to abiotic stress. In addition, it has been suggested that these fungi are able to increase plant disease resistance by activating induced systemic resistance (ISR) . The mode of action of these beneficial fungi in the Trichoderma -plant-pathogen interaction are many, complex and not comple…     

Photoexcited CRY2 interacts with CIB1 to regulate transcription and floral initiation in Arabidopsis

Cryptochromes (CRY) are photolyase-like blue-light receptors that mediate light responses in plants and animals. How plant cryptochromes act in response to blue light is not well understood. We report here the identification and characterization of the Arabidopsis CIB1 (cryptochrome-interacting basic-helix-loop-helix) protein. CIB1 interacts with CRY2 (cryptochrome 2) in a blue light-specific manner in yeast and Arabidopsis cells, and it acts together with additional CIB1-related proteins to promote CRY2-dependent floral initiation. CIB1 binds to G box (CACGTG) in vitro with a higher affinity than its interaction with other E-box elements (CANNTG). However, CIB1 stimulates FT messenger RNA expression, and it interacts with chromatin DNA of the FT gene that possesses various E-box elements except G box. We propose that the blue light-dependent interaction of cryptochrome(s) with CIB1 and CIB1-related proteins represents an early photoreceptor signaling mechanism in plants.     

木霉菌对五种植物病原真菌的重寄生作用

研究了哈茨木霉（Trichoderma harzianum)T88菌株和深绿木霉（T.atroviride）T95菌株对立枯丝核菌（Rhizoctonia solani）、杨树料皮病菌（Valsa sordida）、杨树水泡溃疡病菌（Botryosphaeria ribis）、苹果树腐烂病菌（V.ceratosperma）、贝伦格葡萄座腔菌梨生专化型（B.berengeriana f.sp.piricola）的重寄生作用。对峙培养的结果可观察到，多数情况下，接种后2d内木霉与病原菌接触，随后覆盖或侵入病菌菌落，抑制其生长。光学显微镜和扫描电镜观察结果表明，木霉对不同的植物病原真菌重寄生作用方法不同。可观察到木霉菌缠绕病原菌的菌丝，或沿着病原菌的菌丝平行或波浪式生长，或产生铯状分枝、吸器或附着胞吸附于病原菌的菌丝上，或穿透病原菌的菌丝，最终使病原菌的菌丝细胞原生质浓缩，菌丝断裂等现象。     

G protein signalling involved in host recognition and mycoparasitismrelated chitinase expression in Trichoderma atroviride

Mycoparasitic species of Trichoderma are commercially applied as biological control agents against various fungal pathogens. The mycoparasitic interaction is host specific and includes recognition, attack and subsequent penetration and killing of the host. Investigations on the underlying events revealed that Trichoderma responds to multiple signals from the host (e.g. lectins or other ligands such as low molecular weight components released from the host's cell wall) and host attack is ac…     

3株茶藨生柱锈重寄生木霉菌的形态鉴定及ITS序列分析

[目的]探讨更为客观的木霉菌鉴定手段。[方法]在形态分类的基础上结合分子鉴定手段(rDNA-ITS序列分析),对3株茶藨生柱锈重寄生木霉菌进行分类鉴定。[结果]依据TR1的培养性状和显微特征描述,初步鉴定该菌株为深绿木霉(T.atroviride);TR2、TR3的培养性状和显微特征在一定程度上相似,依据其形态特征,初步鉴定这2个菌株为绿色木霉(T.viridePers.ex Fr.)。从Genbank中深绿木霉和绿色木霉的6个菌株与TR1、TR2T、R3所作的系统发育树可知,TR1应该归为深绿木霉,TR2和TR3属同种真菌,应该归为绿色木霉,这与形态学观察结果一致。[结论]形态特征结合ITS序列分析可作为木霉菌分类鉴定的依据。     

韩国平菇菇床上流行的木霉分离物的形态学和分子生物学分析

自韩国平菇生产地分离到26株木霉(Trichoderma),基于培养特性和菌落形态,这些菌株分为4组,鉴定为Trichoderma sp．K1、Trichoderma sp．K2、T．harizianum,和T．atroviride,其中优势种为Trichoderma sp．K2,其次是Trichoderma sp．K1和T．atroviride。K1和K2不仅在形态上与其它的组有区别,而且相互在生长温度(35℃),菌落形态,分生孢子的形态,分生孢子小梗及其分枝类型,分生孢子的形态等方面有显著差异。对这些菌的ITS、EF-1(和RPB2的序列比较显示出这2个待鉴定的种(K1和K2)不仅与早期报道的木霉种有差异,而且相互间也显著不同。同时发现,EF-1(和RPB2序列比ITS序列在木霉的系统分类上更加客观可靠。基于以上研究结果,我们认为K1和K2是木霉属中的新种,即将给予它们新的名称。     

Factors that contribute to the mycoparasitism stimulus in Trichoderma atroviride strain P1

Trichoderma atroviride strain P1 has been used extensively to study the mycoparasitic mechanisms in the interaction between plant pathogenic host and beneficial antagonistic fungi. Mutants of P1 containing the green fluorescent protein (gfp) or glucose oxidase (gox) reporter systems and different inducible promoters (from the exochitinase nag1 gene, or the endochitinase ech42 gene of P1) were used to determine the factors that activate the biocontrol gene expression cascade in the antagonis…     

拟南芥隐花色素突变体抑制子的筛选及其表型分析

隐花色素（cryptochrome,简称CRY）是与DNA光解酶氨基酸序列高度同源的黄素蛋白,但不具有光解酶活性。拟南芥的隐花色素家族中隐花色素1（CRY1）主要抑制下胚轴伸长,隐花色素2（CRY2）主要调节光周期开花,并且这两个光受体具有部分重叠的功能。cry1cry2双突变体在长日条件下比野生型晚开花。采用激活标签的方法,在cry1cry2双突变体背景下筛选到一个早开花突变体scc17-D (suppressor of cry1cry2#17-Dominant)。TAIL-PCR结果表明,scc17-D突变体的T-DNA插入在第一条染色体上的At1g25440和At1g25450两个基因之间。scc17-D突变体表现出植株矮化、叶片变小、果荚变小和子叶表皮细胞形状发生改变等性状。     

Mechanisms involved in biocontrol and plant induced resistance by richoderma. asperellum (T. harzianum T-203)

Growing awareness of the environmental damage caused by the use of chemical substances for plant disease control in agriculture has raised the need to study biological alternatives, such as activating the defense response of plant crops by inducers not toxic to the environment. Trichoderma spp. are effective biocontrol agents for a number of soilborne pathogens, and are also known for their ability to enhance plant growth and to induce systemic resistance (ISR) in plants. In our laborator…     

紫外光和蓝光调控查尔酮合成酶基因表达的研究

UV-B（280nm～320nm）、UV-A（320nm～390nm）和蓝光（390nm-500nm）经不同光受体和信号传递途径控制植物发育的各个方面。已知的蓝光／UV-A受体有隐花色素CRY1和CRY2，向光性受体有趋光素。氧化还原过程在隐花色素和趋光素信号转导过程中起重要作用。专一的UV-B光受体尚未被鉴定出，存在许多可能的UV-B信号传递途径．拟南芥查尔酮合成酶（CHS）的紫外光和蓝光转录调控成为研究热点。光受体突变体实验表明存在不同的UV-A／蓝光和UV-B光感知系统调控CHS的表达。拟南芥细胞悬浮培养物实验表明UV-B和CRY1信号传递途径在动力学上和药理学上不同。影响诱导CHS转录的启动子元件和转录因子现已被鉴定出。UV-B、隐花色素和光敏色素信号传递途径之间的相互作用调控CHS表达。UV-B途径与UV-A／蓝光途径的协同相互作用使CHS最大量表达。另外，专一的光敏色素经不同的增强和相巨作用正调控CRY1途径，负调控UV-B途径。