BARLEY CULTIVAR IDENTIFICATION BY SDS GRADIENT PAGE ANALYSIS OF HORDEIN

1987 ◽  
Vol 67 (4) ◽  
pp. 927-944 ◽  
Author(s):  
B. A. MARCHYLO
Keyword(s):  
Molecular Weight ◽  
Cultivar Identification ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Resolving Power ◽  
Buffer System ◽  
Barley Cultivar ◽  
Protein Patterns ◽  
Barley Malt ◽  
Barley Cultivars

Sodium dodecyl sulfate gradient polyacrylamide gel electrophoresis (SDSGPAGE) was evaluated to determine whether it could provide improved resolving power of hordein proteins concomitant with improved discrimination (identification) of Canadian barley cultivars. Hordeins were subjected to electrophoresis using a linear gradient gel and a discontinuous buffer system. Proteins were detected using a silver-based color stain and gels were calibrated routinely for molecular weight determinations with a mixture of standard proteins. Molecular weight estimations of hordeins were performed by evaluation of the relationship between log (MW) versus log (R1) and reproducible linear calibration curves and molecular weight estimates for hordeins (CV 0.66%) were obtained. Densitometric analysis of protein band intensities revealed considerable variability in protein staining. Electrophoretic analysis of 100 barley cultivars revealed that, in general, resolution of SDSGPAGE was superior to that of acidic PAGE but considerable duplication of protein patterns among cultivars and their biotypes remained a problem. Cultivar formulae consisting of B, C and D hordein pattern designations were used to prepare a cultivar catalogue of protein patterns. Sixty-five formulae consisting of 30 B, 36 C and 3 D hordein patterns were obtained. SDSGPAGE also was found suitable for barley malt cultivar analysis.Key words: SDS gradient PAGE, barley, cultivar identification, hordein

Variation in goat fibre protein

1989 ◽  
Vol 40 (3) ◽  
pp. 675 ◽  
Author(s):  
DJ Tucker ◽  
AHF Hudson ◽  
A Laudani ◽  
RC Marshall ◽  
DE Rivett
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulfate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Apparent Molecular Weight ◽  
Wool Fibre ◽  
Two Dimensional ◽  
Protein Patterns ◽  
Cashmere Goats

The proteins from a range of cashmere, mohair, angoratcashmere crossbred and wool fibre samples were extracted at pH 8 with 8 M urea containing dithiothreitol, and were then radiolabelled by S-carboxymethylation using iodo(2-14C) acetate. The proteins from each sample were examined by two dimensional polyacrylamide gel electrophoresis in which the separation in the first dimension was according to charge at pH 8.9 and in the second dimension according to apparent molecular weight in the presence of sodium dodecyl sulfate. After electrophoresis the proteins were detected by fluorography. Protein differences in keratin samples from some individual goats existed, although the overall protein patterns were similar. None of the differences were consistent with any one goat fibre type. The protein patterns obtained for fibre samples from individual cashmere goats showed some differences when compared to those found for commercial blends from the same country of origin, indicating that blending can mask any animal-to-animal variation. While the electrophoretic technique does not unequivocally distinguish between cashmere, mohair and angora/cashmere crossbred fibres it does differentiate between wool and goat fibres.

Inheritance of novel high-molecular-weight glutenin subunits in the Tunisian bread wheat 'BT-2288'

Genome ◽  
1988 ◽  
Vol 30 (3) ◽  
pp. 442-445
Author(s):  
R. B. Gupta ◽  
K. W. Shepherd
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Gel Electrophoresis ◽  
Cultivar Identification ◽  
Wheat Cultivar ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Glutenin Subunits ◽  
Bread Making

Using sodium dodecyl sulphate-polyacrylamide gel electrophoresis, three new high-molecular-weight glutenin subunit/subunit combinations were detected in a Tunisian wheat cultivar (BT-2288) and these were designated bands 26, 7 + 11, and 5 + 9. Analysis of 112 testcross seeds revealed that the genes controlling them were additional alleles at Glu-A1, Glu-B1, and Glu-D1 loci, respectively. These alleles enhance the genetic variability available for cultivar identification and possibly for improving the bread-making quality of hexaploid wheat.Key words: Triticum aestivum, Glu-1 loci, high-molecular-weight glutenin subunits.

BARLEY CULTIVAR IDENTIFICATION BY ELECTROPHORETIC ANALYSIS OF HORDEIN PROTEINS: I. EXTRACTION AND SEPARATION OF HORDEIN PROTEINS AND ENVIRONMENTAL EFFECTS ON THE HORDEIN ELECTROPHOREGRAM

1980 ◽  
Vol 60 (4) ◽  
pp. 1343-1350 ◽  
Author(s):  
B. A. MARCHYLO ◽  
D. E. LaBERGE
Keyword(s):  
Gel Electrophoresis ◽  
Room Temperature ◽  
Cultivar Identification ◽  
Polyacrylamide Gel Electrophoresis ◽  
Electrophoretic Analysis ◽  
Western Canada ◽  
Barley Cultivar ◽  
Barley Cultivars ◽  
Extraction And Separation ◽  
Acid Gel

Reproducible hordein electrophoregrams were obtained when hordeins, extracted from Canadian-grown barley cultivars, were subjected to polyacrylamide gel electrophoresis at acid pH (acid-gel PAGE). The hordeins were extracted at room temperature into a solution of 55% (vol/vol) isopropanol containing 2% (vol/vol) monothioglycerol. Extraction of hordein at high temperature (60 °C) produced significant reduction in the number of protein bands available for cultivar identification. Alkylation of the reduced hordeins, prior to acid-gel PAGE, did not improve the resolution or appearance of the hordein electrophoregrams. The effect of environment on the hordein electrophoregram was studied. Four barley cultivars were grown at eight locations in Western Canada during 2 successive years. Hordein electrophoregrams were qualitatively independent of growth location, year of growth and protein content. These results suggest that acid-gel PAGE should prove useful as a technique for barley cultivar identification.

BARLEY CULTIVAR IDENTIFICATION BY ELECTROPHORETIC ANALYSIS OF HORDEIN PROTEINS II. CATALOGUE OF ELECTROPHOREGRAM FORMULAE FOR CANADIAN-GROWN BARLEY CULTIVARS

1981 ◽  
Vol 61 (4) ◽  
pp. 859-870 ◽  
Author(s):  
B. A. MARCHYLO ◽  
D. E. LABERGE
Keyword(s):  
Gel Electrophoresis ◽  
Cultivar Identification ◽  
Polyacrylamide Gel Electrophoresis ◽  
Electrophoretic Analysis ◽  
Polyacrylamide Gel ◽  
Barley Cultivar ◽  
Relative Mobility ◽  
Malting Barley ◽  
Barley Cultivars ◽  
Acid Ph

Hordein proteins, extracted from 62 Canadian-grown two- and six-rowed barley cultivars and six American-grown six-rowed barley cultivars, were separated by polyacrylamide gel electrophoresis at acid pH. Band mobilities were shown to be reproducible within ± 0.005 relative mobility units. Hordein electrophoregram formulae were determined and catalogued for the 68 barley cultivars analyzed. Twenty-five of the cultivars produced unique electrophoregram formulae. The remaining 43 cultivars were divided into 17 groups, each containing between two and eight indistinguishable cultivars. Further analysis of electrophoregram formulae for Canadian-grown barley cultivars revealed that the majority of malting barley cultivars were distinguishable from feed barley cultivars. The usefulness of the acidic PAGE procedure for the identification of Canadian-grown barley cultivars is evaluated.

Heterozygous Abnormal Fibrinogen Osaka III with the Replacement of γ Arginine-275 by Histidine Has an Apparently Higher Molecular Weight γ-Chain Variant

1992 ◽  
Vol 68 (05) ◽  
pp. 534-538 ◽  
Author(s):  
Nobuhiko Yoshida ◽  
Shingi Imaoka ◽  
Hajime Hirata ◽  
Michio Matsuda ◽  
Shinji Asakura
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulfate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Tetraacetic Acid ◽  
Fibrin Monomer ◽  
Sds Page ◽  
Thrombotic Tendency ◽  
Chain Variant

SummaryCongenitally abnormal fibrinogen Osaka III with the replacement of γ Arg-275 by His was found in a 38-year-old female with no bleeding or thrombotic tendency. Release of fibrinopeptide(s) by thrombin or reptilase was normal, but her thrombin or reptilase time in the absence of calcium was markedly prolonged and the polymerization of preformed fibrin monomer which was prepared by the treatment of fibrinogen with thrombin or reptilase was also markedly defective. Propositus' fibrinogen had normal crosslinking abilities of α- and γ-chains. Analysis of fibrinogen chains on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the system of Laemmli only revealed the presence of abnormal γ-chain with an apparently higher molecular weight, the presence of which was more clearly detected with SDS-PAGE of fibrin monomer obtained by thrombin treatment. Purified fragment D1 of fibrinogen Osaka III also seemed to contain an apparently higher molecular weight fragment D1 γ remnant on Laemmli gels, which was digested faster than the normal control by plasmin in the presence of [ethy-lenebis(oxyethylenenitrilo)]tetraacetic acid (EGTA).

Purification and partial characterization of cysteine-glutamate transaminase from rat liver

1977 ◽  
Vol 55 (9) ◽  
pp. 958-964 ◽  
Author(s):  
M. P. C. Ip ◽  
R. J. Thibert ◽  
D. E. Schmidt Jr.
Keyword(s):  
Molecular Weight ◽  
Rat Liver ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Liver Homogenate ◽  
Polyacrylamide Gel ◽  
Gel Chromatography ◽  
Labile Hydrogen ◽  
Apparent Homogeneity

Cysteine-glutamate transaminase (cysteine aminotransferase; EC 2.6.1.3) has been purified 149-fold to an apparent homogeneity giving a specific activity of 2.09 IU per milligram of protein with an overall yield of 15%. The isolation procedures involve the preliminary separation of a crude rat liver homogenate which was submitted sequentially to ammonium sulfate fractionation, TEAE-cellulose column chromatography, ultrafiltration, and isoelectrofocusing. The final product was homogenous when examined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS). A minimal molecular weight of 83 500 was determined by Sephadex gel chromatography. The molecular weight as estimated by polyacrylamide gel electrophoresis in the presence of SDS was 84 000. The purified enzyme exhibited a pH optimum at 8.2 with cysteine and α-ketoglutarate as substrates. The enzyme is inactivated slowly when kept frozen and is completely inactivated if left at room temperature for 1 h. The enzyme does not catalyze the transamination of α-methyl-DL-cysteine, which, when present to a final concentration of 10 mM, exhibits a 23.2% inhibition of transamination of 30 mM of cysteine. The mechanism apparently resembles that of aspartate-glutamate transaminase (EC 2.6.1.1) in which the presence of a labile hydrogen on the alpha-carbon in the substrate is one of the strict requirements.

scholarly journals Isolation and Characterization of Two Proteins fromMoraxella catarrhalis That Bear a Common Epitope

1998 ◽  
Vol 66 (9) ◽  
pp. 4374-4381 ◽  
Author(s):  
John C. McMichael ◽  
Michael J. Fiske ◽  
Ross A. Fredenburg ◽  
Deb N. Chakravarti ◽  
Karl R. VanDerMeid ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Bacterial Attachment ◽  
Vaccine Candidates ◽  
Isolation And Characterization ◽  
Sds Page

ABSTRACT The UspA1 and UspA2 proteins of Moraxella catarrhalisare potential vaccine candidates for preventing disease caused by this organism. We have characterized both proteins and evaluated their vaccine potential using both in vitro and in vivo assays. Both proteins were purified from the O35E isolate by Triton X-100 extraction, followed by ion-exchange and hydroxyapatite chromatography. Analysis of the sequences of internal peptides, prepared by enzymatic and chemical cleavage of the proteins, revealed that UspA1 and UspA2 exhibited distinct structural differences but shared a common sequence including an epitope recognized by the monoclonal antibody 17C7. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), purified UspA1 exhibited a molecular weight of approximately 350,000 when unheated and a molecular weight of 100,000 after being heated for 10 min at 100°C. In contrast, purified UspA2 exhibited an apparent molecular weight of 240,000 by SDS-PAGE that did not change with the length of time of heating. Their sizes as determined by gel filtration were 1,150,000 and 830,000 for UspA1 and UspA2, respectively. Preliminary results indicate the proteins have separate functions in bacterial pathogenesis. Purified UspA1 was found to bind HEp-2 cells, and sera against UspA1, but not against UspA2, blocked binding of the O35E isolate to the HEp-2 cells. UspA1 also bound fibronectin and appears to have a role in bacterial attachment. Purified UspA2, however, did not bind fibronectin but had an affinity for vitronectin. Both proteins elicited bactericidal antibodies in mice to homologous and heterologous disease isolates. Finally, mice immunized with each of the proteins, followed by pulmonary challenge with either the homologous or a heterologous isolate, cleared the bacteria more rapidly than mock-immunized mice. These results suggest that UspA1 and UspA2 serve different virulence functions and that both are promising vaccine candidates.

scholarly journals Characterization of proteins from the cytoskeleton of Giardia lamblia

1983 ◽  
Vol 59 (1) ◽  
pp. 81-103 ◽  
Author(s):  
R. Crossley ◽  
D.V. Holberton
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Giardia Lamblia ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Molecular Size ◽  
Molecular Weights

Proteins from the axonemes and disc cytoskeleton of Giardia lamblia have been examined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis. In addition to tubulin and the 30 X 10(3) molecular weight disc protein, at least 18 minor components copurify with the two major proteins in Triton-insoluble structures. The most prominent minor bands have the apparent molecular weights of 110 X 10(3), 95 X 10(3) and 81 X 10(3). Protein of 30 X 10(3) molecular weight accounts for about 20% of organelle protein on gels. In continuous 25 mM-Tris-glycine buffer it migrates mostly as a close-spaced doublet of polypeptides, which are here given the name giardins. Giardia tubulin and giardin have been purified by gel filtration chromatography in the presence of sodium dodecyl sulphate. Well-separated fractions were obtained that could be further characterized. Both proteins are heterogeneous when examined by isoelectric focusing. Five tubulin chains were detected by PAGE Blue 83 dye-binding after focusing in a broad-range ampholyte gel. Giardin is slightly less acidic than tubulin. On gels it splits into four major and four minor chains with isoelectric points in the pI range from 5.8 to 6.2. The amino acid composition of the giardin fraction has been determined, and compared to Giardia tubulin and a rat brain tubulin standard. Giardins are rich in helix-forming residues, particularly leucine. They have a low content of proline and glycine; therefore they may have extensive alpha-helical regions and be rod-shaped. As integral proteins of disc microribbons, giardins in vivo associate closely with tubulin. The properties of giardins indicate that in a number of respects - molecular size, charge, stoichiometry - their structural interaction with tubulin assemblies will be different from other tubulin-accessory protein copolymers studied in vitro.

Purification and characterization of ferro- and cobalto-chelatases

1988 ◽  
Vol 66 (1) ◽  
pp. 32-39 ◽  
Author(s):  
Eduardo T. Cánepa ◽  
Elena B.C. Llambías
Keyword(s):  
Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Purification And Characterization ◽  
Degree Of Unsaturation ◽  
Apparent Homogeneity ◽  
Optimum Ph ◽  
Single Protein ◽  
Metal Insertion

Pig liver ferrochelatase was purified 465-fold with about 30% yield, to apparent homogeneity, by a procedure involving solubilization from mitochondria, ammonium sulfate fractionation, and Sephacryl S-300 chromatography. The fraction of each purification step had cobaltochelatase as well as ferrochelatase activity. A purified protein of molecular weight 40 000 was found by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. A molecular weight of approximately 240 000 was obtained by Sephacryl S-300 chromatography. Both activities of the purified fraction increased linearly with time until 2 h. but nonlinear plots were obtained with increasing concentrations of protein. Their optimum pH values were similar. Km values were, for ferrochelatase activity, 23.3 μM for the metal and 30.3 μM for mesoporphyrin. and for cobaltochelatase activity. 27 and 45.5 μM, respectively. Fe2+ and Co2+ each protected against inactivation by heat. Pb2+, Zn2+, Cu2+, or Hg2+ inhibited both activities, while Mn2+ slightly activated; Mg2+ had no effect, at the concentrations tested. There appeared to be an involvement of sulfhydryl groups in metal insertion. Lipids, in correlation with their degree of unsaturation, activated both purified activities; phospholipids also had activation effects. We conclude that a single protein catalyzes the insertion of Fe2+ or Co2+ into mesoporphyrin.

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