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SALAMIAH Hajime AKAMATSU Yukitaka FUKUMASA-NAKAI Hiroshi OTANI Motoichiro KODAMA 《Journal of General Plant Pathology》2001,67(2):97-105
The genetic controls of host-specific toxin (HST) biosynthesis and the pathogenicity of A. alternata pathogens have been limited by the asexual nature of the life cycle of these fungi. We used a protoplast fusion system for
A. alternata to analyze the genetics of HST production and its relation to the specific pathogenicity of these pathogens. Drug-resistant
transformants were isolated by genetic transformation, using vectors conferring resistance to hygromycin B and geneticin,
for the A. alternata apple pathotype (AM-toxin producer) and A. alternata tomato pathotype (AAL-toxin producer), respectively. Protoplasts of the respective transformants were fused by electrofusion.
The majority of resultant stable fusants produced both AM- and AAL-toxins and were pathogenic to susceptible cultivars of
both apple and tomato. Pulsed-field gel electrophoresis analysis demonstrated that these fusants (or hybrids) carried small
1.7-and 1.1-Mb chromosomes, characteristic of the parental strains of the apple and tomato pathotypes, respectively. Detection
of the AMT gene, involved in AM-toxin biosynthesis, by polymerase chain reaction revealed that all fusants pathogenic to apple maintained
this gene. Microfluorimetry analysis using propidium iodide staining suggested that the fusants might be diploid.
Received 14 November 2000/ Accepted in revised form 11 December 2000 相似文献
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Yasunori Akagi Masatoki Taga Mikihiro Yamamoto Takashi Tsuge Yukitaka Fukumasa-Nakai Hiroshi Otani Motoichiro Kodama 《Journal of General Plant Pathology》2009,75(2):101-109
To analyze the genetics of host-specific toxin production and its relation to the specific pathogenicity of a mitosporic fungus
Alternaria alternata, we developed a protoplast fusion system. Protoplasts of drug-resistant transformants of the A. alternata tomato pathotype (AAL-toxin producer) and A. alternata strawberry pathotype (AF-toxin producer) were fused by electrofusion. Of five fusion strains examined, two strains were pathogenic
on both tomato and strawberry host plants, whereas the rest of the fusion strains were pathogenic only on tomato. Pulsed-field
gel electrophoresis analysis demonstrated that the hybrid strains pathogenic on both tomato and strawberry carry 1.0- and
1.05-Mb conditionally dispensable (CD) chromosomes derived, respectively, from the parental strains of the tomato and strawberry
pathotypes. On the other hand, the fusion strains appeared to maintain only a single homologous chromosome derived from one
of the parental strain in the case of essential chromosomes (A chromosomes). The results suggest that fusion strains between
two different pathotypes of A. alternata might be haploid resulting from the deletion of extra sets of essential chromosomes in the fused nuclei, whereas the CD chromosomes
derived from each parental strain could be maintained stably in a new genetic background with an expanded range of pathogenicity.
The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank database under the accession numbers AB469331to
AB469354. 相似文献
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Masaki Fukuda Izumi Sendo Satoko Imahori Yukitaka Fukumasa-Nakai 《Journal of Wood Science》2007,53(3):264-267
Electrophoretic analysis of the transmission pattern of mitochondrial plasmids in protoplast cell fusion between compatible
monokaryons of Lentinula edodes indicates that three of the four plasmids carried in parental monokaryons are effectively transferred and replicated in the
protoplast fusants. The two monokaryons, 1158a and 1569a, carried different plasmids that could be distinguished by a single
restriction digest. Electrophoresis of intact plasmids and restriction analyses indicate that all but one of the fusants carry
three of the four possible plasmids, indicating that transmission of plasmids in protoplast fusions is principally biparental
in L. edodes. Thus, heterocytoplasmic cells of L. edodes can be effectively constructed by protoplast cell fusion. In addition, plasmids of the same homology group cannot coexist
in the heteroplasmic cells after protoplast cell fusion.
Contribution No. 382 of the Tottori Mycological Institute 相似文献
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Keisuke Tokimoto Masaki Fukuda Teruyuki Matsumoto Yukitaka Fukumasa-Nakai 《Journal of Wood Science》1998,44(6):469-472
Mycelial protoplasts of two compatible monokaryons ofLentinula edodes were prepared, and electro-fusion was undertaken. Sixteen dikaryons isolated after the electrofusion were tested for their fruiting abilities in comparison to the two dikaryons obtained by reciprocal mycelial mating. There were significant differences in fruiting body yield among the fusants, and some of them produced more fruiting bodies than did normally mated dikaryons. Although these dikaryons were divided into five genetically different groups by isozyme analyses and a cluster analysis, we could not find clear differences among the groups in terms of fruiting body yield, mycelial growth rate, or degree of wood decay. It appeared that fruiting body yield correlated positively with the mycelial growth rate but negatively with the degree of wood decay. 相似文献
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Wiboonsirikul J Kimura Y Kadota M Morita H Tsuno T Adachi S 《Journal of agricultural and food chemistry》2007,55(21):8759-8765
Defatted rice bran was extracted with water and subcritical water at 50-250 degrees C for 5 min. The highest extract yield was achieved at 200 degrees C, at which the maximum amounts of protein and carbohydrate were also obtained. The total phenolic and furfural contents, radical scavenging activity, and antioxidative activity for the autoxidation of linoleic acid increased with increasing treatment temperature. The bran extracts exhibited emulsifying activity except for the extract prepared at 250 degrees C, which was concomitant with the disappearance of its high-molecular-mass substances. The extract prepared at 200 degrees C also had the highest emulsion-stabilizing activity. 相似文献
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SALAMIAH Yukitaka FUKUMASA-NAKAI Hajime AKAMATSU Hiroshi OTANI Keisuke KOHMOTO Motoichiro KODAMA 《Journal of General Plant Pathology》2001,67(1):7-14
Several pathotypes of Alternaria alternata are known to produce host-specific toxins (HSTs) as agents of pathogenicity or virulence. However, investigations into the
genetic controls of HST biosynthesis and pathogenicity of Alternaria pathogens have been limited by the lack of a sexual stage in the life cycle of these pathogens. We report here the development
of a protoplast fusion system and its use for genetic analysis of HST production and specific pathogenicity of the tomato
pathotype of A. alternata that produces AAL-toxin as a HST. Drug-resistant transformants have been isolated by genetic transformation of nonpathogenic
A. alternata (strain O-94) and A. alternata tomato pathotype (strain As-27) with vectors conferring resistance to hygromycin B and geneticin, respectively. Protoplasts
of the respective transformants were fused by polyethylene glycol treatment or electrofusion. Fusion products were selected
by culturing in the presence of both hygromycin B and geneticin, then confirmed by amplification using a polymerase chain
reaction with specific primers to the transforming drug-resistance genes. Stable fusants were purified by successive subcultures
on selective medium and single-spore isolation. The resultant stable fusants, probably inter-strain hybrids, had the same
pathogenicity and toxin production as the wild-type strain As-27. These results suggest that protoplast fusion has potential
applications for genetic analysis of A. alternata pathogens.
Received 8 September 2000/ Accepted in revised form 27 October 2000 相似文献
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Masaki Fukuda Masayoshi Wakayama Masayuki Uchida Yukitaka Fukumasa-Nakai Teruyuki Matsumoto 《Journal of Wood Science》2007,53(4):339-343
Interspecific protoplast fusion between two monokaryotic strains (a methionine auxotrophic and chloramphenicol-resistant Pleurotus ostreatus strain and a wild-type strain of Pleurotus pulmonarius) was carried out to introduce mitochondrial DNA (mtDNA) from P. ostreatus into P. pulmonarius. Because mycelial colonies regenerated on minimum medium containing chloramphenicol only after the treatment of protoplast
electrofusion, the regenerants were regarded as protoplast fusants. The fusants isolated from regenerated colonies were uninucleate,
and their resistance of chloramphenicol seemed to be a stable trait. The fusants mated with P. pulmonarius but not with P. ostreatus, and showed almost identical random amplified polymorphic DNA (RAPD) patterns to that of the parental P. pulmonarius monokaryon. The sizes of mitochondrial genomes were estimated to be 81.5 kb for P. ostreatus monokaryon, 54.9 kb for P. pulmonarius monokaryon, and 84.5 or 86.0 kb for the fusants. BamHI and EcoRI digest patterns of the fusant mtDNAs were almost the same as the parental P. ostreatus monokaryon. From the above results, the fusants seemed to carry nuclei derived from the monokaryon of P. pulmonarius and mtDNA including the chloramphenicol-resistance gene from the P. ostreatus monokaryon, suggesting that the P. ostreatus mtDNA had been introduced into P. pulmonarius cells by interspecific protoplast fusion.
Contribution No. 383 of the Tottori Mycological Institute 相似文献
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