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O L Krasil'nikova L N Anisova N B Romanova Iu E Bartoshevich 《Antibiotiki i khimioterapii͡a》1988,33(7):483-486
Conditions for efficient regeneration in mutant strains of the doxorubicin-producing organism Str. peucetius var. caesius were developed. The effect of the protoplast regeneration on changes in the proportion of the components of the anthracycline complex produced by these strains was shown. Variants with doxorubicin productivity 2 times higher than that of the parent strain were isolated. 相似文献
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D. G. Lyutskanova M. M. Stoilova-Disheva V. T. Peltekova 《Applied Biochemistry and Microbiology》2005,41(2):165-168
Conventional mutagenesis (UV irradiation and exposure to nitrosoguanidine) as well as protoplast formation and regeneration were used to improve the antibiotic activity of a Streptomyces fradiae strain producing tylosin. Variants exceeding the activity of the initial producer strain by 0.5–28.3% were obtained. The most active variants were produced by a combined exposure to UV and nitrosoguanidine, as well as upon regeneration of protoplasts formed from the cells of clones produced by UV irradiation. Unstable inheritance of the trait of increased tylosin production was demonstrated.Translated from Prikladnaya Biokhimiya i Mikrobiologiya, Vol. 41, No. 2, 2005, pp. 189–193.Original Russian Text Copyright © 2005 by Lyutskanova, Stoilova-Disheva, Peltekova. 相似文献
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The tropical agarophyte Gracilaria changii has been much researched and documented by the Algae Research Laboratory, University of Malaya, especially with regards to
its potential as a seaweed bioreactor for valuable compounds. Protoplast regeneration of this seaweed was developed following
the optimization of protoplast isolation protocol. Effect of the concentration and combination of isolating enzymes, incubation
period, temperature, enzyme solution pH, tissue source on the protoplast yields were used to optimize the isolation protocol.
The enzyme mixture with 4% w/v cellulase Onozuka R-10, 2% w/v macerozyme R-10 and 1 unit mL-1 agarase was found to produce the highest yield of protoplast at 28°C and 3 h incubation period. Thallus tips gave higher
yields of protoplasts than middle segments. Freshly isolated G. changii protoplasts were cultured in MES medium. Regeneration of protoplast cell walls after 24 h was confirmed by calcofluor white
M2R staining under UV fluorescence microscopy. The protoplasts with regenerated cell walls then underwent a series of cell
division to produce callus-like cell masses in MES medium. Following this, juvenile plants of G. changii were obtained. 相似文献
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Induction and Characterization of Artificial Diploids from the Haploid Yeast Torulaspora delbrueckii 总被引:2,自引:2,他引:0 下载免费PDF全文
The yeast Torulaspora delbrueckii, which propagates as a haploid, was made into a diploid by treatment with dimethyl sulfoxide (DMSO) on the regeneration of protoplasts. The diploid state was stably inherited; the cell volume was three times that of the parent strain and the cellular DNA content was two times that of the parental strain. No essential difference was found between diploids induced by DMSO and those formed through intraspecific protoplast fusion. The diploid strains sporulated fairly well, with their cells converting directly into asci. Random spore analysis revealed that diploids induced through protoplast fusion gave rise to auxotrophic segregants (haploids) with the parental genetic marker or to segregants formed by recombination, while diploids induced by DMSO from a doubly auxotrophic parent gave rise to no recombinant, indicating that it was chromosomally homoallelic in nature. The magnesium level in the protoplast regeneration medium was found to be an important factor for inducing diploid formation. At 0.2 mM magnesium diploids appeared even in the absence of DMSO, while at 2 mM magnesium diploids never appeared unless DMSO was added to the regeneration medium. Evidence is provided that the diploids induced by DMSO or a low magnesium level are due to direct diploidization but not protoplast fusion. UV light irradiation of intact cells (without protoplasts), 10% of which survived, also produced diploids among this surviving population. From these results we conclude that the perturbation of protoplast regeneration or of cell division by the treatments mentioned above somehow induced direct diploidization of T. delbrueckii. 相似文献
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Jari Vehmaanperä 《FEMS microbiology letters》1988,49(1):101-105
Abstract A method for efficient polyethylene glycol (PEG)-mediated transformation of Bacillus amyloliquefaciens protoplasts with plasmid DNA is described. The best conditions found for protoplast regeneration included using 0.45 M sucrose both during the cultivation of the cells and (as an osmotic stabilizer) during their treatment with lysozyme, whereas 0.25 M sodium-succinate was added to the regeneration plates. Under these conditions about 5–10% of input cells regenerated. The highest transformation frequency with plasmid DNA was obtained with a PEG 6000 concentration of 22.5% (w/v). Transforming B. amyloliquefaciens strains with the plasmid pUB110 isolated from B. amyloliquefaciens resulted in 2–4 · 105 transformants/μg DNA, 100–1 000-times as high as with DNA from Bacillus subtilis , suggesting a restriction barrier between the two species. Transformation of B. amyloliquefaciens with plasmids pC194 or pE194 cop -6 gave poor yields and no restriction barrier could be demonstrated for these plasmids. However, by curing pC194 from one of the transformants, a mutant strain compatible to both the plasmids could be isolated, yielding 2–3·104 transformants/μg DNA. Both laboratory and industrial B. amyloliquefaciens strains could be transformed with the procedure. 相似文献
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Satoru Furukawa Tomoki Azuma Toshihide Nakanishi Masahiro Sugimoto 《Applied microbiology and biotechnology》1988,29(2-3):248-252
Summary
Corynebacterium glutamicum R-18 is a strain forl-isoleucine production. Polyethylene glycol (PEG)-induced protoplast fusion was applied to improve the strain of thisl-isoleucine producer. Strain R-18 was fused with anl-lysine producerC. glutamicum S-37, becausel-isoleucine andl-lysine are synthesized from a common intermediate, aspartate--semialdehyde. Two thousand fusants were checked for their phenotypes. Most of the fusants accumulatedl-lysine, and only 0.9% of the fusants accumulatedl-isoleucine. Two strains, F-28 and F-91, were selected and cultivated in production medium. Fusant F-28 accumulated 12.1 g/l ofl-isoleucine and 4.8 g/l ofl-lysine, and fusant F-91 accumulated 4.8 g/l ofl-isoleucine and 13.0 g/l ofl-lysine, while the parental strains R-18 and S-37 accumulated 9.5 g/l ofl-isoleucine and 26.8 g/l ofl-lysine, respectively. Sugar consumption activity was improved by protoplast fusion, and thel-isoleucine production rate of F-28 was 2.4 times higher than that of R-18. 相似文献
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桃褐腐病菌(Monilia fructigena)原生质体制备及再生条件 总被引:3,自引:0,他引:3
以桃褐腐病菌(Monilia fructigena)为供试菌株,研究了酶系组成、液体培养基、菌龄、酶解温度、酶解时间对原生质体制备的影响,以及等渗液、固体再生培养基、酶解时间对原生质体再生的影响。结果表明:Fries(1/2)液体培养基培养24h,在10mg/mL崩溃酶+5mg/mL纤维素酶+20mg/mL蜗牛酶+10mg/mL溶菌酶的混合酶液中28°C酶解4h为桃褐腐病菌原生质体制备的最佳条件。采用液体再生涂布平板法,以含Ca2+的STC为等渗液的液体培养基和含蔗糖及Ca2+的Fries(1/2)固体培养基为桃褐腐病菌原生质体再生的最佳条件。经过观察与测定,再生菌株保持了原有的培养性状和致病性,接种桃果实后发病率为100%。 相似文献
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本研究在初步实现水稻原生质体培养的程序化后,选用普通栽培稻P339和特种稻苏御糯选的原生质体为融合亲本,利用碘乙酸(IA)和罗丹明-6G(R-6G)这两个代谢互补抑制剂钝化处理亲本原生质体,确定了合适的抑制条件。P339用0.25mmol/L IA,苏御糯选用50μg/ml R-6G分别经30min钝化处理,通过PEG和高Ca~(2 )、高pH法诱导融合,异源融合体具有代谢互补效应,经培养得到愈伤组织17块,并进一步分化获得不同形态的再生植株12棵。移栽存活的再生植株成熟后可育,通过对这些植株的形态以及酯酶和过氧化物酶同工酶电泳的分析表明是融合后的体细胞杂种植株。 相似文献
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Liutskanova DG Stoilova-Disheva MM Peltekova VT 《Prikladnaia biokhimiia i mikrobiologiia》2005,41(2):189-193
Conventional mutagenesis (UV irradiation and exposure to nitrosoguanidine) were used to produce and regenerate protoplasts, aiming at increasing the antibiotic activity of a Streptomycesfradiae strain producing tylosin. Variants exceeding the activity of the initial producer strain by 0.5-28.3% were obtained. The most active variants were produced by a combined exposure to UV and nitrosoguanidine, as well as upon regeneration of protoplasts formed from the cells of clones produced by UV irradiation. Unstable inheritance of the trait of increased tylosin production was demonstrated. 相似文献
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The influence of protoplasting and protoplast regeneration in the presence of polyethylene glycol on antibiotic activity, components of antibiotic complexes and antibiotic resistance in Streptomyces hygroscopicus 155 was studied. It was shown that the protoplasting and protoplast regeneration influenced the antibiotic activity. The protoplast fusion resulted in increased isolation of variants with higher antibiotic activity. The processes also affected the components of the antibiotic complexes but had no effect on the strain resistance to some antibiotics. 相似文献
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In order to genotype hybrid genomes of distant asymmetric somatic hybrids, we synthesized hybrid calli and plants via PEG-mediated
protoplast fusion between recipient tall fescue (Festuca. arundinacea Schreb.) and donor wheat (Triticum aestivum L.). Seventeen and 25 putative hybrid clones were produced from the fusion combinations I and II, each with the donor wheat
protoplast treated by UV light for 30 s and 1 min, respectively. Isozyme and RAPD profiles confirmed that ten hybrid clones
were obtained from combination I and 19 from combination II. Out of the 29 hybrids, 12 regenerated hybrid plants with tall
fescue phenotype. Composition and methylation-variation of the nuclear and cytoplasmic genomes of some hybrids, either with
or without regenerative ability, were compared by genomic in situ hybridization, restriction fragment length polymorphism,
and DNA methylation-sensitive amplification polymorphism. Our results indicated that these selected hybrids all contained
introgressed nuclear and cytoplasmic DNA as well as obvious methylation variations compared to both parents. However, there
were no differences either in nuclear/cytoplasmic DNA or methylation degree between the regenerable and non-regenerable hybrid
clones. We conclude that both regeneration complementation and genetic material balance are crucial for hybrid plant regeneration. 相似文献
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大球盖菇原生质体再生及单核化特性的研究* 总被引:1,自引:0,他引:1
大球盖菇原生质体再生条件及单核化特性结果。原生质体再生速度极快,涂布平板3d后肉眼即可见明显的再生菌落形成,在PGPM再生培养基上再生率为0.97~2.0%,渗稳剂种类对再生率无明显影响,但可影响再生菌落形态,液体预培养1~2d,再生率明显下降;大球盖菇原生质体单核化率高达77.6%,且再生双核体和再生单核体在形成再生菌落时无时间差,其生长速度亦无快慢之分,液体预培养可显著减少单核化率,再生单核体中存在亲本两种交配型,但二者的比率不为1。 相似文献
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E. de Vries-Uijtewaal L. J. W. Gilissen E. Flipse K. Sree Ramulu W. J. Stiekema B. de Groot 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1989,78(2):185-193
Summary
Agrobacterium transformation of stem internodes of four monohaploid (839-79, 849-7, 851-23, 855-1) and two diploid (M9 and HH260) potato genotypes using hairy root-inducing single (LBA 1020, LBA 9365, LBA 9402) and binary (LBA 1060KG) vectors is reported. Various media and successive culture steps were tested for plant regeneration from different transformed root clones. The fate of introduced genetic markers in root clones and regenerated plants (hairy root phenotype, hormone autotrophy, opine production, kanamycin resistance, -glucuronidase activity), the ploidy stability and protoplast yield were analysed. The transformation efficiency of stem internodes (hairy root production) and the regeneration capacity of the transformed root clones greatly differed within and between the various potato genotypes. The regenerated plants obtained after transformation with both types of vectors often showed the absence of one or more genetic markers. However, transformation with the binary Agrobacterium vector generally resulted in the stable presence of the opines in all transformed root clones and most regenerated plants. In HH260, transformation efficiency, plant regeneration of transformed root clones, protoplast yield and ploidy stability were the highest as compared to the other genotypes. The application of these transformed plants as marker lines in gene mapping and gene expression studies is indicated. 相似文献
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Dr. M. C. Liechty S. G. Bradley K. L. Kasweck 《Journal of industrial microbiology & biotechnology》1987,2(2):71-77
Summary Fusion and regeneration of protoplasts ofNocardia asteroides strains ATCC 3318, IMRU W3599 and HIK B971 have been used to study genetic recombination in this species. Protoplasts were produced by treatment with lysozyme, following incubation with glycine. Mutants of ATCC 3318 were grown in peptone yeast extract medium at 32°C prior to protoplast production to maximize protoplast frequency, whereas mutants of IMRU W3599 and HIK B971 were grown in trypticase-soy broth. Glycine concentrations favoring protoplast formation varied from 1.5% to 5% depending on strain. For all strains, protoplast formation was complete 1 h after addition of 5 mg/ml lysozyme. Protoplasts were fused by addition of 50% polyethylene glycol-1000. In general, 25% of the protoplasts could be regenerated. The incidence of recombinant recovery was increased up to 750-fold. The distribution of recombinant phenotypes in matings was similar for protoplast fusion and conventional crosses. 相似文献
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Methods were developed for the isolation of large numbers of healthy protoplasts from two species of the agarophyte Gracilaria; G. tikvahiae McLachlan and G. lemaneiformis (Bory) Weber-van Bosse. This is the first report of protoplast isolation and cell division in a commercially important, phycocolloid-producing red seaweed, as well as for a member of the Florideophycidae. The optimal enzyme composition for cell wall digestion and protoplast viability consisted of 3% Onozuka R-10, 3% Macerozyme R-10, 1% agarase and 0.5% Pectolyase Y- 23 dissolved in a 60% seawater osmoticum containing 1.0 M mannitol. The complete removal of the cell wall was confirmed by several different methods, including electron microscopic examination, and the absence of Calcofluor White (for cellulose) and TBO (for sulfated polysaccharide) staining. Spontaneous protoplast fusion was observed on several occasions. Protoplast viability was dependent upon the strain and age of the parent material, as well as the mannitol concentration of the enzyme osmoticum. Cell wall regeneration generally occurred in 2-6 days; cell division in 5-10 days. Protoplast-produced cell masses up to the 16-32 cell stage have been grown in culture. However, efforts to regenerate whole plants have been unsuccessful to date. 相似文献