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  • 1
    Electronic Resource
    Electronic Resource
    Rapid isolation of plant peroxidase. Purification of peroxidase a from Petunia (1984)
    Oxford, UK : Blackwell Publishing Ltd
    Physiologia plantarum 60 (1984), S. 0 
    Details
    ISSN: 1399-3054
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: A rapid isolation procedure was developed for purification of peroxidase a from Petunia hybrida. Rapid isolation was possible since about 15% of the extracellular protein from stem tissue obtained by vacuum infiltration followed by centrifugation of the tissue represents peroxidase. Purification of peroxidase a from intercellular fluid was achieved by two acetone precipitation steps followed by DEAE-cellulose chromatography.Three different forms of peroxidase were eluted from DEAE-cellulose at different NaCl concentrations. Isoelectric focusing showed, however, a pI of 3.8 for all three forms of peroxidase a. Only part of the peroxidase a enzymes bound to Concanavalin A indicating heterogeneity in the carbohydrate part. Homology of peroxidase a to the peroxidase G1 group from tobacco is discussed.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1399-3054.1984.tb06066.x
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  • 2
    Electronic Resource
    Electronic Resource
    Cellular location of peroxidase isoenzymes in leaf tissue of Petunia and their affinity for Concanavalin A-Sepharose (1985)
    Springer
    Planta 164 (1985), S. 89-95 
    Details
    ISSN: 1432-2048
    Keywords: Concanavalin A ; Leaf (peroxidases) ; Peroxidase isoenzymes ; Petunia (peroxidases)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The cellular location of three peroxidase isoenzymes (PRX) in mature leaf tissue of Petunia and their affinity for Concanavalin A-Sepharose were investigated. The isoenzymes PRXa, PRXb and PRXc were identified by their positions in starch-gel zymograms. The fast-moving anodic and cathodic peroxidase bands, the isoenzymes PRXa and PRXc respectively, were the most active peroxidases in extracellular extracts. The molecular forms of PRXa showed a tissue-specific distribution between midrib and remaining leaf tissue. An intermediate-moving anodic peroxidase band, the isoenzyme PRXb, was the most active peroxidase released after extraction of isolated mesophyll protoplasts. Small amounts of the peroxidase isoenzymes were present in cell-wall-bound fractions. Incubation of a crude protein fraction with Concanavalin A-Sepharose showed that the isoenzyme PRXb bound more firmly to Concanavalin A-Sepharose than the isoenzymes PRXa and PRXc, of which only one molecular form bound partly. The results are discussed with respect to a possible function of one of the peroxidase isoenzymes, and a possible role of oligosaccharide chains in determining the cellular location of plant peroxidases is suggested.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00391030
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  • 3
    Electronic Resource
    Electronic Resource
    Inbred testing of tomato (Lycopersicon esculentum L.) F1 varieties by ultrathin-layer isoelectric focusing of seed protein (1990)
    Weinheim : Wiley-Blackwell
    Electrophoresis 11 (1990), S. 824-829 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: To test seed lots of tomato F1 hybrid varieties for the presence of undesirable inbred seeds by electrophoresis, a method has been developed on the basis of ultrathin-layer isoelectric focusing. The method is based on the genetic variation of the seed protein PRS-1 which could be visualized by isoelectric focusing of a 5 mm NaCl-soluble seed protein extract in a pH 6-9 gel followed by protein staining. Two genetic variants of the PRS-1 protein, PRS-1+ and PRS-11, were found among open-pollinated varieties, as well as among F1 hybrid varieties. The isoelectric points (pI) of the PRS-1 proteins are 7.1 and 6.1 for PRS-1+ and PRS-11, respectively. The PRS-1 protein is unique to seed tissue and is located primarily in the embryo. A genetic 1:2:1 segregation of the gene Prs-1 among several F2 populations shows monogenic inheritance. Analysis of commercial F1 hybrid varieties from several seed companies indicated that the Prs-11 allele, in contrast to the Prs-1+ allele, is primarily present in gene pools of „Moneymaker type“ tomatoes. The described method is generally applicable to all tomato F1 varieties that are heterozygous for the gene Prs-1. With the described method one person can routinely analyze more than 768 seeds per day.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150111009
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  • 4
    Electronic Resource
    Electronic Resource
    Horizontal ultrathin-layer multi-zonal electrophoresis of DNA: An efficient tool for large-scale polymerase chain reaction (PCR) fragment analysis (1997)
    Weinheim : Wiley-Blackwell
    Electrophoresis 18 (1997), S. 2861-2864 
    Details
    ISSN: 0173-0835
    Keywords: DNA electrophoresis ; Polymerase chain reaction fragments ; Large-scale analysis ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: A rapid and efficient procedure for large-scale analysis of polymerase chain reaction (PCR) fragments in the range of 200-3000 base pairs is presented. The procedure is based on horizontal ultrathin-layer multi-zonal (HUME) electrophoresis of PCR fragments in polyacrylamide gels followed by silver staining. HUME gels can be prepared rapidly using a simple procedure called the flap technique. The electrophoretic set-up allows the use of multi-channel pipettes for sample loading. Separation and detection of the PCR fragments from sample preparation to silver staining can be carried out in 2 h. Using four electrophoresis units, one technician can analyze 400 PCR fragment samples in 2 h.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150181524
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  • 5
    Electronic Resource
    Electronic Resource
    Polymorphism of pepper (Capsicum annuum L.) seed proteins studied by two-dimensional electrophoresis with immobilized pH gradients: Methodical and genetic aspects (1994)
    Weinheim : Wiley-Blackwell
    Electrophoresis 15 (1994), S. 297-304 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: A population of ten morphologically similar inbred lines of pepper (Capsicum annuum L.) has been investigated for polymorphism of seed proteins by two-dimensional (2-D) electrophoresis with immobilized pH gradients. To reveal as much variation as possible, both the water- and the urea/detergent-soluble protein fraction were electrophoretically analyzed and subsequently visualized by silver staining. The 2-D patterns were subjected to computer analysis to be able to establish genetic variation. A high number of the seed proteins were found to be variable as to presence/absence: these were 68 out of 184 reproducible water-soluble proteins and 34 out of 419 reproducible urea/detergent-soluble proteins. Comparison of the 2-D data of the water-soluble and the urea/detergent-soluble proteins, which represent the biggest part of all extractable seed proteins, showed that both protein fractions have proteins in common, but the variable proteins found in both fractions were non-identical. The difference of variability scored in both solubility fractions was discussed. Genetic distances between all pairs of inbred pepper lines were calculated and a genetic tree was constructed. A correlation analysis was carried out to correct for genetic linkage and for secondary modifications, to have a more proper estimate of genetic distances. In both cases the dendrograms showed two distinct genetic groups of five inbred lines. This electrophoretic study was done in order to utilize the genetic distance data in breeding for heterosis. The genetic distance data presented will be used to validate the assumption that there is a higher chance to achieve better hybrid performance when the genetic distance between the parents is as great as possible. From the fact that we found a high level of genetic variability within a population of ten morphologically similar inbred lines, it can be concluded that 2-D electrophoresis can be efficiently applied in pepper breeding.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150150150
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  • 6
    Electronic Resource
    Electronic Resource
    The terminator, an apparatus for simultaneous homogenization of 96 small seeds individually (1992)
    Weinheim : Wiley-Blackwell
    Electrophoresis 13 (1992), S. 880-881 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: An apparatus was developed to homogenize 96 individual seeds of tomato, pepper, cabbages, and other seeds of similar size in a 96-well tissue culture plate in 45 s. The apparatus, named the terminator, consists of two parts: a base plate that can hold the 96-well tissue culture plate, and a vibrating part that is attached to a variable transformer. The seeds are homogenized by the vibrating action of 96 small homogenizers that are attached to the vibrating part and that fit perfectly into the wells of the tissue culture plate. The seed homogenates made in this way can be used for electrophoretic analysis of genetic variations of seed proteins.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.11501301192
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  • 7
    Electronic Resource
    Electronic Resource
    Genetic variability of pepper (Capsicum annuum L.) seed proteins studied by 2-D electrophoresis with immobilized pH gradients (1992)
    Weinheim : Wiley-Blackwell
    Electrophoresis 13 (1992), S. 774-777 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Ten pepper (Capsicum annuum L.) inbred lines were successfully differentiated by two-dimensional electrophoresis with immobilized pH gradients. Qualitative polymorphism of water-soluble and urea/detergent-soluble seed proteins, respectively, was investigated by computer analysis and used for establishing a dendrogram derived from maximum-parsimony analysis. The dendrogram calculated from urea/detergent-soluble proteins shows four types of distance indices, whereas water-soluble proteins show two sets of inbred lines with similar intraset distance indices. The validity of the dendrograms with respect to quantitative inherited traits, such as cold tolerance and earliness, will be tested by field trials.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.11501301168
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  • 8
    Electronic Resource
    Electronic Resource
    State of the art of large-scale genetic purity testing of hybrid vegetable seeds using isoelectric focusing at PetoSluis (1996)
    Weinheim : Wiley-Blackwell
    Electrophoresis 17 (1996), S. 502-504 
    Details
    ISSN: 0173-0835
    Keywords: Vegetables ; Isoelectric focusing ; Genetic purity-testing ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: During the past ten years we have been engaged in developing and applying isoelectric focusing techniques to test the genetic quality of vegetable seeds. We started with isoelectric focusing using carrier ampholytes (IEF-CA), and continued research with isoelectric focusing in immobilized pH gradients (IEF-IPG) and two-dimensional electrophoresis (IPG-DALT). In addition, we have developed equipment and procedures for large-scale seed and seedling homogenization, sample preparation and semi-automatic gel staining. Moreover, we have optimized the sample application and gel running setup for large-scale analysis. We have developed hybrid purity (inbred) testing methods for all important vegetables, e.g., melon, cole crops, tomato, pepper, watermelon, squash, cucumber, radish etc. using either IEF-CA or IEF-IPG of seed or seedling proteins, followed by specific or general protein staining. To indicate the efficiency of the equipment and procedures developed we present results of two of our hybrid purity test methods, namely for brassica using polymorphism for phosphoglucomutase (PGM) from dry seeds, and for tomato using polymorphism for alcohol dehydrogenase (ADH) from imbibed seeds. We show that one person can routinely analyze 1536 individual seeds per day at a cost of about US $0.11 per seed for chemicals, materials, and electrophoresis equipment.
    Additional Material: 2 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150170313
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  • 9
    Electronic Resource
    Electronic Resource
    Isoelectric focusing in the vegetable seed industry (1998)
    Weinheim : Wiley-Blackwell
    Electrophoresis 19 (1998), S. 1780-1787 
    Details
    ISSN: 0173-0835
    Keywords: Isoelectric focusing ; Vegetable seeds ; Hybrid purity ; Large-scale testing ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Different types of electrophoretic procedures were developed for hybrid purity testing based on starch gel electrophoresis (SGE), vertical polyacrylamide gel electrophoresis (PAGE), and isoelectric focusing (IEF). For the most important vegetables these methods are much faster than plant grow-outs and relatively inexpensive. Compared to SGE and PAGE methods, horizontal IEF proved to be more efficient for large-scale hybrid purity testing. These developments were made possible by the basic work of Harry Rilbe and improvements that were initiated as a result of Rilbe's work. The present paper describes a number of milestones during this developmental period, starting with the isoelectric focusing concept of Harry Rilbe up to the large-scale application of IEF. Further, a comparison of IEF with DNA fingerprinting methods along with the future of both techniques is discussed with respect to hybrid purity testing in the vegetable seed industry. When it comes to a choice between the use of either IEF or a DNA-based method, efficiency and efficacy determines the method which is best suited for hybrid purity testing. It is also concluded that in the future we will see an increased use of both IEF as well as DNA-based methods for hybrid purity testing because expectations of growers has increased; consequently they will accept fewer inbreds in a hybrid variety, especially when growing in a greenhouse.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150191040
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  • 10
    Electronic Resource
    Electronic Resource
    A rapid and economical method for hybrid purity testing of tomato (Lycopersicon esculentum L.) F1 hybrids using ultrathin-layer isoelectric focusing of alcohol dehydrogenase variants from seeds (1991)
    Weinheim : Wiley-Blackwell
    Electrophoresis 12 (1991), S. 64-69 
    Details
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: In the cultivated tomato (Lycopersicon esculentum L.), two electrophoretic variants of the enzyme alcohol dehydrogenase (ADH) encoded by the alleles Adh-1+ and Adh-11 are found. A rapid and economic method for testing the hybrid purity of tomato F 1 seeds, based on the expected presence of Adh-1 alleles, was developed. The method is based on the analysis of the ADH variants by ultrathin-layer isoelectric focusing, pH range 3-10, of crude extracts from imbibed seeds followed by enzyme activity staining. The isoelectric points (pIS) of the ADH variants were estimated to be 5.5 and 5.7 for Adh-1+ and Adh-11; respectively. Using the procedure described and a newly developed sample applicator strip, it is Possible for one person to routinely analyze 1152 seeds per day using only a single electrophoresis unit. An investigation of a large number of inbred lines, both experimental and commercial hybrids, together with open-pollinated varieties, showed the potential of the method. Among F1 hybrids, a higher frequency of the Adh-11 allele was found than among open pollinated varieties, suggesting that F1 hybrid breeding has resulted in a higher frequency of Adh-11 alleles by selection of linked genes.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/elps.1150120112
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