Summary The ascomycete fungus Ascochyta rabiei, an important pathogen of the grain legume crop chickpea (Cicer arietinum L.) in the Mediterranean region, has not been adequately characterized in molecular terms. We therefore used DNA fingerprinting, with synthetic oligodeoxynucleotides complementary to simple repetitive sequences, to pathotype different isolates of the fungus. Six single-spored A. rabiei isolates were first categorized using a host differential set of nine chickpea genotypes. Seedlings were inoculated under controlled environmental conditions, and disease severity was recorded 9 days after inoculation. DNA was extracted from in vitro-grown mycelia of the six purified fungal isolates, restricted with EcoRI, HinfI, MboII and TaqI, and fingerprinted with radiolabeled (GATA)4, (GTG)5, (CA)8, and (TCC)5, respectively. High levels of polymorphism were detected with optimal enzyme/probe combinations that allow one to discriminate between the isolates. The potential of DNA fingerprinting with simple repetitive sequences can thus be expanded to the identification of fungal races and pathotypes. The characterization of the geographic distribution and genetic variability of pathotypes will facilitate the selection of suitable host cultivars to be grown in specific regions. 相似文献
Cochhobolus heterostrophus has alternate genes (MAT-1 andMAT-2) at its mating-type locus. Transformants of aMAT-1 or aMAT-2 strain carrying a transgene of opposite mating type can self and are dual maters; the transgene, however, promotes development of pseudothecia only, not ascospores. To determine if the resident gene interferes with the function of the transgene, transformation vectors were designed to delete different amounts (2.5 kb, 5.7 kb, and 6.3 kb) of DNA at theMAT locus. Deletions occurred at a higher frequency (about 90% of transformants) with linearized plasmid than with circular plasmid (about 15% of transformants), and all three vectors were equally efficient at gene replacement. BothMAT-1 andMAT-2 could be deleted with the same set of vectors. Re-transformation of deletion strains (regardless of deletion size) with a wild-type copy ofMAT restored full mating ability, indicating that the residentMAT gene interferes with function of theMAT transgene. Moreover, sexual development was normal whether theMAT transgene integrated at the homologous or at an ectopic site. 相似文献
We have established stable, bright green fluorescent protein (GFP)- or red fluorescent protein (RFP)-expressing HT-1080 human
fibrosarcoma clones. These cell lines showed similar cell proliferation rates and high-frequency experimental lung metastasis.
The HT-1080-GFP and -RFP clones enable simultaneous real-time dual-color imaging in the live animal. HT-1080 cells were transduced
with retroviral vectors containing GFP or RFP and the neomycin resistance gene. Stable transformants were selected stepwise
with G418 up to 800 μl/ml. Subsequently, high GFP- or RFP-expressing clones, HT-1080-GFP or HT-1080-RFP, respectively, were
selected. 3×106 cells from each clone were mixed and injected into the tail vein of SCID mice. The cells seeded the lung at high frequency
with subsequent formation of pure green and pure red colonies as well as mixed yellow colonies with different patterns visualized
directly on excised lungs. The lung metastases were also visualized by external fluorescence imaging in live animals through
skin-flap windows over the chest wall. Lung metastases were observed on the lung surface of all mice. SCID mice well tolerated
multiple surgical procedures for direct-view imaging via skin-flap windows. Real-time metastatic growth of the two different
colored clones in the same lung was externally imaged with resolution and quantification of green, red, or yellow colonies
in live animals. The color coding enabled determination of whether the colonies grew clonally or were seeded as a mixture
with one cell type eventually dominating, or whether the colonies grew as a mixture. The simultaneous real-time dual-color
imaging of metastatic colonies described in this report gives rise to the possibility of color-coded imaging of clones of
cancer cells carrying various forms of gene of interest.
This revised version was published online in July 2006 with corrections to the Cover Date. 相似文献
Institute of Pharmacology, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. V. Val'dman.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 107, No. 4, pp. 431–433, April, 1989. 相似文献
Introduction: Fungal diseases are a threat to human health. Therapies targeting the fungus continue to lead to disappointing results. Strategies targeting the host response represent unexplored opportunities for innovative treatments. To do so rationally requires the identification and neat delineation of critical mechanistic pathways that underpin human antifungal immunity. The study of humans with single-gene defects of the immune system, i.e. inborn errors of immunity (IEIs), provides a foundation for these paradigms.
Areas covered: A systematic literature search in PubMed, Scopus, and abstracts of international congresses was performed to review the history of genetic resistance/susceptibility to fungi and identify IEIs associated with fungal diseases. Immunologic mechanisms from relevant IEIs were integrated with current definitions and understandings of mycoses to establish a framework to map out critical immunobiological pathways of human antifungal immunity.
Expert opinion: Specific immune responses non-redundantly govern susceptibility to their corresponding mycoses. Defining these molecular pathways will guide the development of host-directed immunotherapies that precisely target distinct fungal diseases. These findings will pave the way for novel strategies in the treatment of these devastating infections. 相似文献
Biolistic (biological ballistic) and protoplast-mediated procedures were compared as methods for transforming strains of Gliocladium virens and Trichoderma harzianum. For biolistic transformation, conidia were bombarded using a helium-driven biolistic device to accelerate M5 tungsten particles coated with plasmid or genomic DNA. DNA from either source contained a bacterial hygromycin B resistance gene (hygB) as a dominant selectable marker. The same sources of DNA were also used to transform protoplasts using a standard polyethylene glycol-CaCl2 protoplast fusion protocol. Hygromycin B-resistant (HygBR) transformants were recovered from all strains, methods, and DNA sources except for genomic DNA used with the protoplast method. The biolistic procedure was technically simpler, and increased transformation frequency and genetic stability in the progeny as compared with the protoplast-mediated transformation. Southern analysis of homokaryotic HygBR progenies showed that the transforming sequences were integrated into the genome of the recipient strains, and apparently were methylated. This is the first study presenting detailed results on biolistic transformation of a filamentous fungus. 相似文献
The projection of muscle afferent fibres to the medulla oblongata and upper spinal cord was studied in the cat by using transganglionic transport of wheat germ agglutinin-horseradish peroxidase conjugate. The results demonstrate a precise, musculotopic termination pattern in the external cuneate nucleus; thus, fibres from the intrinsic muscles of the paw terminate medially; those from forearm, arm, and shoulder muscles terminate progressively more laterally; and those from neck and thoracic muscles terminate in the ventrolateral and dorsolateral parts, respectively. Muscle afferent fibres to the main cuneate nucleus terminate in the ventral "reticular" region of the nucleus, with a sparse projection also to the ventral part of the rostral and caudal regions, including the base of the dorsal horn. Fibres from the neck muscles terminate slightly more laterally in the ventral region than do those from the limb muscles, but otherwise, and thus contrary to the case in the external cuneate nucleus, no topographic organization was detected. In the spinal cord, projection was found to laminae I and V, and from the musculature of the back of the neck to the central cervical nucleus. 相似文献