Acid Phosphatase Zymograms Research Articles

Inherited chilling tolerance in somatic hybrids of transgenic Hibiscus rosa-sinensis x transgenic Lavatera thuringiaca selected by double-antibiotic resistance

Improvement of Hibiscus rosa-sinensis for increased frost tolerance has been attempted through somatic hybridization with the frost tolerant Lavatera thuringiaca. Cell suspensions from Hibiscus and Lavatera were transformed with A. tumefaciens harboring plasmids containing selectable genes coding for kanamycin and hygromycin resistance, respectively. We provided evidence that H. rosa-sinensis and L. thuringiaca were transformed by strong selection of transformed calluses in medium containing antibiotics, by GUS activity determination in protein extracts and by molecular confirmation of chromosomal integration and expression of the selectable genes. Protoplasts isolated from a kanamycinresistant Hibiscus callus and from a hygromycin-resistant Lavatera callus were fused and selected in medium containing both antibiotics. We determined unambiguously that the regenerated double-antibiotic resistant clones obtained are indeed somatic hybrids through analysis of acid phosphatase zymograms and nuclear DNA content. Plant regeneration through somatic embryogenesis was accomplished from both isolated protoplasts and transgenic calluses of L. thuringiaca. However, regeneration from the double-antibiotic resistant fusant calluses was unsuccessful. Analysis of the somatic hybrids at the callus level showed that chilling and freezing tolerance are governed by independent genetic components. The somatic hybrids displayed significant improvement for chilling tolerance at conditions lethal to H. rosa-sinensis, although frost tolerance was not expressed.

Identification of Phytophthora nicotianae on the aerial portion of eucalypt seedlings

AbstractThe pathogenicity, physiology and biochemical features of three isolates of Phytophthora. nicotianae var. nicotianae from Eucalyptus delegatensis and E. globulus seedlings were investigated. The acid phosphatase zymograms and total protein profiles confirmed the specific identity of the 3 isolates. Despite significant differences in mycelial growth and zoosporangium production, no differences were noted in pathogenicity. Artificial inoculations revealed a variability in resistance to P. nicotianae within Eucalyptus species.

Identification of Shiitake strains and some other basidiomycetes : Protein profile, esterase and acid phosphatase zymograms as an aid in taxonomy

Lentinula edodes strains exhibited a typical total protein pattern when native soluble proteins were separated by isoelectric focusing at a pH gradient of 3·5–10. Intraspecies variation was found when using a narrower acidic pH gradient of 4–6. Esterase zymograms showed variation between the shiitake strains. However, the variation was wider between different wood-rotting species. Although the acid phosphatase isoenzyme patterns were all similar in the shiitake strains, variation was observed between the different basidiomycete species. The use of acid phosphatase and esterase as an aid in the taxonomy of wood rotting basidiomycetes is discussed.

Isoenzyme and Total Protein Analysis by Agarose Isoelectric Focusing, and Taxonomy of the Genus Acanthamoeba

ABSTRACTThirty axenically grown reference strains belonging to 15 different Acanthamoeba spp. were investigated for isoenzyme patterns by agarose isoelectric focusing in the pH range 3–10. Zymograms of acid phosphatase, leucine amino peptidase, malate dehydrogenase, propionyl esterase, glucose phosphate isomerase, phosphoglucomutase, and alcohol dehydrogenase were compared. The same strains were also analyzed for protein patterns separated by agarose isoelectric focusing in a pH gradient of 5–8. The results suggested changes in taxonomy within morphology group II of Pussard & Pons. Acanthamoeba paradivionensis becomes a synonym of A. divionensis. Although this species seems to be related to A. rhysodes, it could not be concluded that the species names are synonyms since the type strain of A. rhysodes was not available for comparison. In the subgroup A. polyphaga–A. quina–A. lugdunensis, A. lugdunensis becomes the species name for pathogenic strains of this subroup, A. quina for the nonpathogenic strains, while A. polyphaga is the species name for an atypical strain. Two strains of A. castellanii showed different zymograms from strain Neff of this species, but related protein patterns. In group III, A. pustulosa is found to be a synonym of A. palestinensis, while one strain of A. lenticulata is also found to belong to the A. palestinensis species. All other species names in both morphology groups could be retained as valuable, on the basis of the techniques used. Group I was not investigated, as axenic cultures could not be obtained.

Genetic analysis and geographical distribution of acid phosphatase isozyme in cultivated rice, Oryza sativa L.

Of 3, 746 rice varieties and strains collected from all over the world, 2, 842 (75.87%), 900 (24.03%), 2 (0.05%) and 2 (0.05%) showed J-, I-, O- and M-type zymograms of acid phosphatase, respectively. The gene symbols Acp19 Acp2Nul, Acp1-4 Acp2Fa, Acp1Nul Acp2Fa and Acp1Nul Acp2Nul were given for the J-, I-, O- and M-type zymograms, respectively, following the format described by Pai et al. (1975). Hereafter, however, in this paper the authors mainly used only the gene symbol of the Acp1 except the M-type zymogram, because the Acp1 locus was closely linked with Acp2 as described by Pai et al. (1975). F2 generations obtained from the F1 between the I- and J-types gave a good fit to a 1:2:1 ratio suggesting that these zymograms were controlled by Acp1 locus which carried two codominant alleles. The O-type zymogram was spontaneously induced mutant from the I-type, and controlled by a single recessive silent allele, Acp1Nul. The M-type was artificially induced mutant from the J-type, and governed by double recessive alleles, Acp1Nul Acp2Nul. From the results described above, all varieties and strains of the sativa rice plants were divided into either the genotypes Acp19/Acp19 or Acp1-4/Acp1-4. The gene Acp1-4 is distributed in the low latitude area with high frequencies, while the gene Acp19 predominates in the high latitude area.

Preliminary Study of the Isoenzymes of Ostracoda

Esterases and acid phosphatases were studied of 6 species of ostracod genera Cyprinotus Br. and Notodromas Lillj. from Poland, using method of electrophoretic separation of isoenzymes on polyacrylamide gel (no zymograms could be obtained of the peroxidase and dehydrogenase isoenzymes). For separations a plate device was applied, which enabled to perform accurate comparisons of the pictures obtained. There were about 20 specimens of given species in a sample, thus zymograms of mentioned proteins resultant of studied population were examined. Dependence of the esterase and acid phosphatase zymograms on ontogenetic development, sex, species and morphological variation within the species was analysed. Esterases appear to be more inert than the acid phosphatases (they are markedly less strongly differentiated than acid phosphatases ; the conclusions on similarity between the species as determined by the former coincide with the conclusions from the morphological studies, otherwise than in the case of the latter). Usefulness of results obtained is discussed, and necessity of further studies is proved.

Temperature Regulation of Germination in Crimson Clover Seeds

Seeds of Dixie crimson clover (Trifolium incarnatum cv. Dixie) completed germination in 36 hours at 20 C. At 10 C germination was delayed by 24 hours. At 30 C only 20% germinated and the rest remained viable for a long time but not germinable. Different patterns of adenylate energy state and zymograms of acid phosphatase and esterase were observed from seeds grown under the three temperatures for 24 hours. Varied specific activities of protease, alpha-amylase, ATPase, RNase, acid phosphatase, glutamine synthetase, and fumarase were also found. Protein-synthesizing ability was proportional to temperature. These data indicate that temperature regulates seed germination at multiple sites.

GENIC ANALYSIS FOR ACID PHOSPHATASE ISOZYMES INORYZA PERENNISANDO.SATIVA

It was concluded from genetic experiments that various acid phosphatase zymograms observed among the rice strains belonging to Oryza perennis Moench (wild) and O. sativa L. (cultivated) were controlled by alleles at three loci, Acp1, Acp2, and Acp3. At Acp1seven codominant alleles were found each producing a group of three major (A, M, and C) and three minor (a, m, and c) bands on starch gel. The alleles determined mobility of the band group. Statistical analysis of the data taken from densitometric tracings of zymograms indicated that hybrids between strains with different mobilities would produce three major and three minor hybrid bands each occurring between two corresponding parental bands. In addition, a null form lacking all these bands was found that had a recessive allele, Acp1Nu1. The Acp1locus was independent of Px1(specifying peroxidase OC, 2A, and 4A bands), but was linked withPx2(specifying peroxidase 4C band). The Acp2locus carried two codominant alleles specifying bands Fa and Sa, respectively, and a recessive silent allele, Acp2Nu1. This locus was closely linked with Acp1, yet it was distinguished from the latter as different allelic combinations were observed among parental strains. At Acp3alleles specifying bands B and b were found which were peculiar to O. perennis subsp. barthii; other perennis and sativa strains appeared to have their recessive silent alleles.

Electrophoretic characterization of six selected enzymes of peanut cultivars

The isozyme patterns (both anodic and cathodic) of esterase, catalase, leucine aminopeptidase, acid phosphatase, alcohol dehydrogenase and INT oxidase in individual seeds from several peanut cultivars ( Arachis hypogaea) were characterized by polyacrylamide and starch gel electrophoresis in relation to the stages of seed development, maturity, and germination, geographic areas where grown and phylogenetic relationship. Of the six enzymes examined, only esterase contained cathodic isozymes, of which the patterns served to distinguish between the Spanish and the Virginia-type peanuts. Anodic esterase and acid phosphatase zymograms of early developing and germinating peanuts could be distinguished from those of predormant and mature seeds and the latter showed much intravarietal variation which was consistent among cultivars and the geographic areas where grown. Anodic isozymes of catalase, leucine aminopeptidase, alcohol dehydrogenase and INT oxidase were synthesized very early in peanut development and remained constant through maturity and to at least 24 hr germination; they were consistent within and between peanut cultivars and they were not influenced by the environmental conditions of the areas where the peanuts were grown. The consistency of the isozyme patterns within and between cultivars supports the suggestion that plant breeding programs used to develop superior cultivars have produced genetic uniformity in peanuts.

GENETIC CONVERGENCE OF THE SPECIFIC ACID PHOSPHATASE ZYMOGRAMS IN <i>ORYZA SATIVA</i>

In a number of rice cultivars, Oryza sativa, acid phosphatase zymograms were found to be of two kinds. Genetic experiments revealed that the isozymic difference between the two was due to alleles at Acp1 and Acp2 loci which are closely linked. Those two zymograms were detected in wild rice strains of O. perennis which is believed to be the wild progenitor of O. sativa. The genetic behavior of the isozymes was very similar to that of the isozymes of O. sativa. Although O. perennis showed many different acid phosphatase zymograms, semi-wild perennis-sativa strains were found like O. sativa to have the same two kinds. Probably, domestication, even in a primitive stage, may lead to genetic convergence for the genes specifying the acid phosphatase zymograms of rice.

Studies of acid phosphatase and nonspecific esterase activities in rat adrenal glands following operative stress.

Acid phosphatase and esterase activities were studied in adrenal glands obtained from rats killed at regular intervals following surgical stress (cauda equina transection). Zymograms of acid phosphatase produced by starch gel electrophoresis revealed increased reactivity in the operated samples. With esterases, a slightly different pattern was observed in the operated group, which exhibited a few additional bands particularly in the cathode region. This was confirmed by densitometric analysis of the gel strips. Two of these additional bands were organophosphate-sensitive and the remaining few were activated by p-chloromercuribenzoate. These latter bands appeared to arise from splitting of the preexisting organophosphate-resistant bands present in control zymograms. Biochemical assay of the two hydrolytic enzymes demonstrated a remarkable similarity in their responses to operative stress—probably implying a general lysosomal activation. Both enzymes exhibited a peak activity 8 hr after operation, followed by a gradual decline. Both organophosphate-sensitive and organophosphate-resistant esterases contributed toward the rise in total esterase activity. Histochemical studies on tissue sections revealed a more reactive adrenal cortex in the operated group, but were of little help in localizing the additional esterase activity observed in gel strips.