Technique Of Starch-gel Electrophoresis Research Articles

Gordon Henry Dixon. 25 March 1930—24 July 2016

Gordon H. Dixon was a highly influential scientist who excelled in all the stages of his career and in each laboratory of every institution at which he was a member. Gordon began his graduate career at the University of Cambridge and completed his studies at the University of Toronto on the subject of transpeptidation reactions in biological systems. At an early stage of his career, he developed the technique of starch gel electrophoresis with Oliver Smithies and made important discoveries on the structure of human haptoglobins. Subsequently, Gordon contributed to the determination of the structure and active sites and mechanisms of action of trypsin and chymotrypsin and made seminal discoveries related to understanding the structure of the protein hormone insulin. However, he is best known for his later studies on the regulation of protamine genes and chromatin transitions in spermatogenesis. He is often considered to be the father of the protamine molecular biology underlying this gene system and his research is continually cited to this day. Gordon contributed to the identification and sequencing of the protamine genes, the discovery that methionine was required for initiation of protamine synthesis, understanding the roles of histone hyperacetylation and protamine phosphorylation, and the generation of the highly compact nucleoprotamine structure present in sperm, primarily using trout as a model system. Gordon is fondly remembered as a brilliant and very generous scientist by all his mentees, representing all levels from undergraduates to PhD and postdoctoral fellows, who provided a scientifically stimulating atmosphere in which they could develop their careers. From a more personal perspective, he also had a fun side and was devoted to his family. Gordon was a person who, owing to his legacy of great science and his fundamental humanity, still lives in the memory of many people.

Allozyme variations in six natural populations of scots pine (Pinus sylvestris) in Turkey

Genetic variation in six natural populations of Scots pine (Pinus sylvestris L.) was determined with isoenzyme analyses. For this purpose, haploid female gametophytes of seeds and horizontal starch gel electrophoresis technique were used. A total of 17 loci and 58 alleles were observed in studying 10 enzyme systems. The average proportion of polymorphic loci for populations ranged from 58.8% to 70.6%. The average number of alleles per locus per population was 2.65. The mean estimated expected heterozygosity (He) of populations was 0.294. A rather high proportion of genetic diversity (96.4%) was due to within-population variation and the remaining (3.6%) was due to variation among populations. The level of gene flow (Nem) was found to be 6.69 per generation. Nei’s genetic distance coefficient ranged from 0.006 to 0.027 (mean 0.017) among all possible population pairs. The mean value of Nei’s genetic distance is similar to the values reported for other European Scots pine populations. The low mean value of Nei’s genetic distance among populations is enough to explain low interpopulation variation. According to genetic variation parameters, three out of six populations (Akdagmadeni-Yozgat, Refahiye-Erzincan and Vezirkopru-Samsun) appear to be preferable populations for genetic conservation and forest tree breeding programs.

Genotypes, allele frequencies and dynamic distribution on resistance-associated esterase genes of Culex pipiens complex in Hangzhou

To investigate the genotypes , allele frequencies and dynamic distribution on resistance associated esterase genes of Culex pipiens complex in Hangzhou. The PCR-restriction fragment length polymorphism (PCR-RFLP) assay was applied to type the resistance associated esterase genes, and dynamic surveillance on frequencies of the resistance associated esterase gene of natural population of Culex pipiens complex in Hangzhou during 2003-2005, and phenotype of the resistance associated esterase genes were detected by esterase starch gel electrophoresis technique. The PCR-RFLP assay of esterase allele genes for three consecutive years disclosed four esterase genotypes, namely, the world-wide highly active homozygous Est beta 1(1) (50%-54%), homozygous Est beta 2 (29%-34%), heterozygous Est beta 1(1)/beta 2 (5%-10%) and Est beta N (3.13%) of a new homozygous genotype. The research of the resistance associated esterase genes phenotype in natural population of Culex pipiens complex in Hangzhou in 2005 with esterase starch gel electrophoresis technique revealed four major types, namely, Est beta 1(1) (61%), Est alpha 2/beta 2 (12%), Est alpha 8/beta 8 (7%) and sensitive phenotype (29%). There should be various resistance associated esterase genotypes in natural population of Culex pipiens complex in Hangzhou. During the period of 2003-2005, Est beta 1(1) was the major type; Est alpha 2/beta 2 was the second. Est beta N was a new esterase genotype detected in 2005 only with a mere percentage of 3.13%. As for its resistance to the new insecticide, a follow-up study should be needed. The molecular typing of the amplified esterase gene should be consistent with the resistance associated esterase genes phenotype.

Isozyme electrophoretic characterization of 29 related cultivars of lily (Lilium spp.)

AbstractIsozyme banding patterns (IBPs) were studied for cultivars of lily (Lilium spp.) by means of horizontal starch‐gel electrophoresis (SGE). An array of continuous histidine‐citrate buffer systems at eight ranges of pH and four extraction buffers were tested. On the basis of this survey, the extraction buffer two (Eb‐2) and the buffer system E at pH 7.7 were found to be suitable for detection of lily isozymes. Using the SGE technique, IBP in catalase (CAT; EC 1.11.1.6), esterase (EST; EC 3.1.1.1), malate dehydrogenase (MDH; EC 1.1.1.37), malic enzyme (MAL; EC 1.1.1.40), peroxidase (POX; EC 1.11.1.7), phosphoglucomutase (PGM; EC 2.7.5.1), phosphoglucose isomerase (PGI; EC 5.3.1.9) and 6‐phosphogluconate dehydrogenase (PGD; EC 1.1.1.44) were assayed. In total 29 cultivars were tested in this study: nine were analysed for all eight enzyme systems, 16 cultivars for seven systems, three for six, and one for five enzyme systems. Some IBP were identified as section‐specific biochemical markers. Eight enzymes systems were analysed by constructing a dendogram using the unweighted pair group method, arithmetic average (UPGMA) cluster analysis. The analysis indicated that the lily cultivars could be separated from other Lilium species, except for two L. x formonlogi cultivars:‘Hakuba’ and ‘Hakuko’ which could not be distinguished from each other by the isozyme patterns assayed here. This study shows that isozymes can provide useful biochemical markers for lily cultivar identification and to estimate the phylogenetic relationships among those cultivars.

Allozymic variability in Spanish populations ofCeratitis capitata

Introduction. The Tephritidae Ceratitis capitata is one of the most important agricultural pests in the world. Eradication programmes need as much genetic information as possible to be efficient. It is vital to know the genetic variation that exists in the areas thought to have been important in the expansion of the species, such as the Iberian Peninsula. The aim of this study was to determine the genetic structure of Spanish C. capitata populations and the relationships between them. Materials and methods. We studied the genetic varia- bility of populations of C. capitata (eight wild samples and one laboratory strain) through the use of the horizontal starch gel electrophoresis technique and the assay of fifteen enzyme- coding loci chosen at random. Results. The quantity of variability detected in this polypha- gous species was not high. Of the 15 loci studied, only four were clearly polymorphic. No significant differences were found in any comparison of populations collected from different hosts. Discussion. The distribution patterns of this variation seem to be the result of gene flow and selection in the form of agricultural practices. Spain / Ceratitis capitata / electrophoresis / genetic variation

Esterases no exame da estrutura populacional de Camu-camu (Myrciaria dubia (Kunth) McVaugh-Myrtaceae)

Dois sistemas enzimáticos (esterase e esterase-D), analisados pela técnica de eletroforese em gel de amido, em folhas jovens de plantas cultivadas em terra firme, de sementes provenientes de três amostras de populações naturais de camu-camu, Myrciaria dubia (Kunth) McVaugh-Myrtaceae, procedentes de Iquitos, Boa Vista e Uatumã, revelaram a presença de 6 locos: Est-1, Est-2, Est-3, Est-4, Est-D1 e Est-D2. Dois dos seis locos gênicos examinados no presente estudo (Est-3 e Est-D2) mostraram-se polimórficos, sendo desse modo considerados valiosos no estudo de caracterização da estrutura populacional da espécie. Os padrões de polimorfismo revelados nos locos Est-3 e Est-D2 de camu-camu, são típicos de enzimas monoméricas e diméricas, respectivamente. O loco Est-3 apresentou um grande desbalanço genético dentro e entre as amostras populacionais examinadas, devido ao excessivo número observado de plantas heterozigóticas em relação ao número esperado. O loco Est-D2 apresentou um polimorfismo exclusivo para os alelos Est-D2¹,Est-D2² e Est-D2³, e um bom balanço genético na amostra populacional de Uatumã. Em função disso, dentre os demais locos gênicos aqui investigados, o loco Est-D2 parece ser o mais adequado para identificação e delimitação de prováveis estoques de camu-camu. Portanto, recomenda-se que esse loco esteja presente na lista dos marcadores isoenzimáticos a serem usados em futuras prospecções sobre genética populacional dessa espécie na região amazônica. Dados sobre a distribuição das freqüências alélicas, estimativas das distâncias genéticas, e estimativas de variação genética nos 6 locos de esterases examinados, foram eficazes na demonstração de diferenças genéticas entre as amostras populacionais examinadas da espécie. Os maiores valores de heterozigozidade média (0,1353); proporção de locos polimórficos (0,33) e número médio de alelos por loco (1,33) revelados na amostra populacional de Uatumã, devem-se ao fato dessa amostra ter apresentado um loco polimórfico a mais (Est-D2), em relação as demais amostras investigadas.

Análise da variabilidade de isoenzimas em acessos e cultivares de girassol

O número de genótipos de girassol (Helianthus annuus L.) disponíveis aos agricultores é pequeno e pouco se conhece sobre os genótipos provenientes de outros países que possam ser utilizados no melhoramento genético desta cultura. O objetivo deste trabalho foi analisar a variabilidade genética de 31 acessos e de cinco cultivares nos sistemas isoenzimáticos fosfoglicomutase (PGM), 6fosfogliconato desidrogenase (PGD), fosfoglicoisomerase (PGI) e esterase (EST). Utilizou-se a técnica de eletroforese em gel de amido/penetrose para o fracionamento das isoenzimas da PGM, PGI e PGD e focalização isoelétrica para EST. Foram detectados um total de seis locos e 14 alelos para estes quatro sistemas. Observou-se variantes alélicas para os locos Pgi2, Pgm1, Pgd2, cada um com dois alelos, e para Est1, que apresentou seis alelos. Os testes de adequação aos modelos de equilíbrio de Hardy-Weinberg e de Wright evidenciaram que em dez acessos houve um desvio significativo de endocruzamento.

Reference range of serum haptoglobin is haptoglobin phenotype-dependent in blacks

Reference values for serum haptoglobin (Hp), were established in a Black Zimbabwean population. The upper limit (2.15 g/l) is comparable to the one in Caucasians, but the lower limit (0.12 g/l) is much lower than the proposed interim international reference limit (0.3 g/l). Subjects that typed as Hp 0-0 by starch gel electrophoresis technique were retyped using high performance gel permeation chromatography. This resulted in a 32% decrease in the frequency of Hp 0-0, but an increase in Hp 2-2 and Hp 2-1M phenotype frequencies. In the Zimbabwean Blacks, the Hp 0-0 frequency was estimated to be 2.9%. Haptoglobin reference values were found to be Hp phenotype-dependent; highest values were found in Hp 1-1 (median 0.88 g/l; range 0.31–1.69 g/l) and in Hp 2-1 (median 0.90 g/l; range 0.31–2.22 g/l) and lower values (median 0.66 g/l; range 0.13–1.79 g/l) in Hp 2-2 subjects. The Hp 2-1M phenotype was characterized by low reference values (0.18–1.25 g/l) ( P<0.05). In three cases of the rare variant Hp Johnson, high Hp concentrations were found (median 1.57 g/l; range 0.98–1.57 g/l).

Biochemical polymorphism in yellow catfish, Mystus nemurus (C&V), from Thailand.

Yellow catfish, Mystus nemurus (Cuv. & Val.), is becoming one of the major freshwater species farmed by aquaculturists in Southeast Asia. It was of interest to examine levels of genetic subpopulation differentiation among samples of this species obtained from parts of its range, as well as to compare the genetics of wild and hatchery-bred fish. Horizontal starch gel electrophoresis and histochemical staining techniques were used to examine genetic variation within and among eight wild and one hatchery populations of M. nemurus from northern, northeastern, central and southern Thailand. Four tissues (heart, liver, kidney, and muscle) from individual specimens were used to analyze variations at 23 protein-coding loci. Fifteen of the 23 loci examined (65.22%), namely, ACP*, AAT-1*, EST-1*, EST-2*, GPI*, IDH-1*, IDH-2*, MDH-1*, MDH-2*, MDH-3*, ME*, PGM*, 6PGD*, SOD*, and HB*, were polymorphic at the 0.95 level. Observed heterozygosities ranged from 0.041 to 0.111, with an average of 0.068 +/- 0.028. Genetic distances ranged from 0.005 to 0.164. The greatest genetic distance was found between the Chainat and the Suratthani populations (0.164), a level indicative of subspecific differentiation in M. nemurus from within Thailand.

Biochemical characterization of Tunisian grapevine varieties

&lt;p style="text-align: justify;"&gt;61 tunisian autochton grapevine varieties have been collected for biochemical identification. Isozymes analysis with starch gel electrophoresis technique was used to confirn or to cancel random denominations awarded to the majority of these local varieties. In our conditions, concentrated plant extracts were obtained from vigorous donnant canes newly cut off from selected mother plants during automn. These allowed us to dispose of rigorously interpretable isozyme banding patterns of GPI and PGM systems and to overcome difficulties often related to the use of PGM system. The study of GPII and PGM enzyme systems allowed us to classify the autochton accessions into 16 different groups from which 5 groups containing only 2 or 3 varieties.&lt;/p&gt;&lt;p style="text-align: justify;"&gt;On the other hand, the study of AAT and peroxydase enzyme systems has shown stable and legible isozyme banding patterns allowing to discriminate between equivalent accessions such as Sakasly and Kahli (two black local vines very similar), 3 varieties of Bidh Hamem (Bidh Hamem, Bidh Hamem Rafraf and Bidh Hamem Sfax), and 2 varieties of Bezzoul Kelba Bidha (Sfax and Gabes). In addition, certain varieties having for longtime the same denominations were characterized. A case of point the 4 varieties Khalt meaning mixture (Bouchemma, Abiedh, Mdaouer and Souche 1) and the 3 varieties of Arich (Ahmar, Dressée, and Jerba) were proved to be completely different from each other. In the same way, Bezzoul Khadem has been differed from Hemri variety. The complementary use of berry colour allowed to discriminate between Saouadi, Khdhiri and Jebbi varieties and to subdivise the remainig groups into sub-groups.&lt;/p&gt;&lt;p style="text-align: justify;"&gt;The study of GPI, PGM, AAT and peroxydase isozyme banding patterns in combination with berry colour has led to establish a classification of the 61 autochton varieties into 37 groups including 26 varieties definitely differentiated through the results of this biochemical study.&lt;/p&gt;

Chromosomal location of eight isozyme loci in rice using primary trisomics and monosomic alien addition lines

Starch-gel electrophoresis and isoelectric focusing techniques were used to detect polymorphism and chromosomal location of isozyme loci in rice (Oryza sativa L.). Polymorphism for four new isozyme loci- Mdh-3, Gdh-2, Fk-1, and Fk-2- was detected. Primary trisomic analysis showed that Dia-1 and Fk-1 are located on chromosome 1, Mdh-1 on chromosome 5 and Fk-1, and Gdh-2 and Tpi-1 as a cluster of genes on chromosome 11. Monosomic alien addition line analysis revealed that Mdh-3 and Pgd-2 are located on chromosomes 5 and 6, respectively.

Isozyme markers and genetic variability in three species of Centrosema (Leguminosae)

Isozyme patterns and their genetic control in three Centrosema species are described. Seven isozymatic systems (aspartate aminotransferase, glucose-6-phosphate isomerase, phosphoglucomutase, anodal peroxidase, malate dehydrogenase, 6-phosphogluconate dehydrogenase, and isocitrate dehydrogenase) were studied in 18 populations and several breeding lines of C. acutifolium, C. brasilianum and C. pubescens, using starch gel electrophoresis techniques. All systems, except glucose-6-phosphate isomerase, are described for the first time in these species. A total of 17 isozyme loci were scored; this represents the largest set of Mendelian loci known up to now in Centrosema species. Isozyme polymorphism and variability within and between populations and species were relatively high and allowed discrimination among species

BIOCHEMICAL SYSTEMATICS AND EVOLUTION OF THE CRAYFISH GENUS ORCONECTES (DECAPODA: CAMBARIDAE)

Phylogenetic relationships among 30 species of the North American crayfish genus Orconectes were investigated by examining 56 presumptive enzyme-coding loci using the technique of horizontal starch gel electrophoresis. Specimens were obtained from each of the 10 recognized subgenera and represent 14 of the 15 species-groups. Specimens from 2 additional genera (2 species of Cambarus and 3 species of Procambarus) were also included as outgroups. The most striking result in this investigation is the clustering of the troglobitic species of the sub-genus O. (Orconectes) in a group by themselves. The allozyme data indicate that these cave species are very divergent (Nei's D ≥ 0.3577) from other members of the genus and appear more closely related to the outgroup species of Cambarus (mean D = 0.3522). This result suggests that O. (Orconectes) is a monophyletic lineage that deserves generic recognition, as previously proposed by Creaser (1962). In addition, no monophyletic lineages were found that support the other subgenera or ally their species groups, a result consistent with recent DNA sequence data. However, a bootstrap analysis of these data suggests that the allozyme phylogeny is poorly resolved among the other subgenera of Orconectes. This indicates, perhaps, that the limits of the allozyme technique to distinguish phylogenetic relationships at the subgeneric level were reached (i.e., insufficient levels of variation were detected). The distribution of alleles at some loci, however, might provide insight into the evolutionary history and past dispersal patterns of these crayfish species. These data lend support to the thesis of Fitzpatrick (1987) that some ancestral orconectid stocks established themselves west of the Mississippi River where they eventually diverged from the stocks found to the east on the Cumberland Plateau. Unique alleles found only in species of the Ozark Highlands suggest also that this area played a pivotal role in the evolution and diversification of ancestral crayfish stocks.

Genetic Variation in Natural Populations of the Large Cranberry, Vaccinium macrocarpon Ait. (Ericaceae)

1816, the large cranberry has become a regionally important small fruit crop in the United States. In order to assess genetic variation in naturally occurring populations of the large cranberry, sampling was conducted at ten sites distributed from Wisconsin east to Massachusetts, and south to Delaware. Starch gel electrophoresis and substrate-specific staining techniques were used to resolve 23 putative Mendelian loci. Population data collected as individual genotypes were analyzed, generating measures of population structure and genetic diversity. All populations were found to exhibit low levels of genetic variation, e.g., expected heterozygosity (Hexp 0.00 + 0.002-0.057 + 0.030). Relative to other plant species, total genetic diversity was extremely low (H, 0.048) with the majority of all genetic variation due to differences among individuals within populations. This genetic homogeneity was corroborated by Nei's (1978) genetic identity which ranged from 0.977 to one. Recent events in the evolutionary history of this taxon are proposed to have played an important role in determining population genetic structure in this species.

Population genetics of the shrimp Artemesia longinaris (crustacea, penaeidae) on the south-west atlantic coast

1. 1. In order to test the hypothesis of the existence of two subpopulations of the shrimp Artemesia longinaris on the south-west Atlantic coast, as was suggested by Nascimento (Nascimento P. A. M., 1983; Naturalia, Sao Paulo, 8, 33–47), a gene frequency analysis of four A. longinaris subpopulations was done by starch-gel electrophoresis techniques. 2. 2. Samples were collected along the Rio Grande (Brazil) and Mar del Plata (Argentine) coasts. 3. 3. Three of the eight enzyme systems analyzed were polymorphic (MDH, PGI and IDH). 4. 4. No significant allele frequency differences were found among samples from the putative subpopulations. 5. 5. The contingency Chi-square test ( P > 0.112), the low values of D m (0.0007), G st (0.0039) and F st (0.008), and the high genetic identity ( I = 0.999) demonstrated the large homogeneity among A. longinaris samples. 6. 6. It is concluded, thus, that there is no genetic evidence to support the existence of two subpopulations of A. longinaris on the south-west Atlantic coast.

Comparison of Isozyme Polymorphism in Races ofCochliobolus carbonum

Differences in isozyme polymorphisms between races of Cochliobolus carbonum were compared using starch gel electrophoresis techniques. Of 36 enzymes analyzed, isozyme polymorphisms were detected for aspartate aminotransferase, esterase, glucose-phosphate isomerase, leucine aminopeptidase, and mannitol dehydrogenase. Specific combinations of polymorphisms were found to be diagnostic of race pathotype for the isolates used in this study (...)

Errata: Molecular Studies of Thamnophiine Snakes: 1. The Phylogeny of the Genus Nerodia

Errata: Molecular Studies of Thamnophiine Snakes: 1. The Phylogeny of the Genus Nerodia

Sagartia troglodytes (Anthozoa: Actiniaria) consists of two species

The sagartiid sea anemone Sagartia troglodytes (Price) has usually been considered to consist of 2 morphological varieties h k e d by intermediate forms (and therefore presumably by interbreeding). The techniques of starch gel electrophoresis and chromosome preparation, combined with an analysis of reproductive biology, were employed to investigate populations around the SW coasts of Britain to test this assumption. Var. decorata is extremely variable both morphologically and genetically with high levels of polymorphism and heterozygosity (P = 61 %, H, = 0.27). It reproduces exclusively by out-crossing, oviparous means producing maximal genotypic diversity and good gene flow between populations (Nei's Genetic Identity, 1 = 0.99). Var. ornata is less variable morphologically but genetically diverse (P = 62 %, H. = 0.47) due to 'fixed heterozygosity' at some locl, which is maintained by extensive clonal reproduction within populations producing relatively low inter-population identity (f = 0.86). Var. decorata has a chromosonle number of approximately 24 whereas var. ornata has approximately 48. On this basis it is proposed that var.decorata should retain the species name S. troglodytes (Price) and that var. ornata be restored to species status as Sagartia ornata (Holdsworth, in Gosse 1860).

Allozyme variation in the milkweed butterflies (Lepidoptera: Danainae)

Seventeen enzyme systems, representing 22 zones of activity, are surveyed in 20 species of Danainae (some represented by multiple samples) and one species of Acraeinae, by the technique of horizontal starch gel electrophoresis. Descriptions of each of the enzyme systems is given, including a brief review of their use in previous lepidopteran electrophoretic investigations. Each currently recognized danaine taxon is considered from the viewpoint of whether or not it can be characterized using allozymes. Enzyme polymorphism in relation to average heterozygosity and population structure is discussed, as are the possible reasons for the observed allozymic differences between the N American and Australian populations of the monarch, Danaus plexippus. Finally, the potential for allozymic data in the classification of the Danainae is examined and it is concluded that such data are likely to be of most use at the generic level and below.

AN ELECTROPHORETIC STUDY OF EVOLUTION IN CAPSICUM (SOLANACEAE).

Most recent treatments of the peppers, genus Capsicum (Solanaceae), recognize four or five domesticated taxa and 20 or more wild species (Eshbaugh, 1980a). The distribution of the species is fairly well documented with the exception of the Brazilian taxa. The genus includes a number of wide ranging species as well as several narrow endemics, e.g., C. galapagoensis, C. cardenasii, etc. All taxa are at least facultative self-fertilizers with the exception of C. cardenasii. A survey of chromosome numbers indicates n = 12 or 13 (Pickersgill, 1977), suggesting two lines of evolution which may eventually require some realignment at the generic level. Changing concepts on evolution of the genus have led from the hypothesis of C. frutescens as the single wild progenitor of all the domesticates (Davenport, 1970; Ramalingam, 1972; Jett, 1973), to four or five distinct progenitors each giving rise to a domesticate (Heiser et al., 1971) and alternatively, to three independent lines of evolution leading to domesticated C. baccatum, C. pubescens, and a complex of C. annuum, C. chinense, and C. frutescens (Eshbaugh, 1980b). Various approaches to investigating and understanding evolution in the genus have included comparative morphological investigation, karyotype analysis (Ohta, 1962; Shopova, 1966; Pickersgill, 1977) and the study of breeding behavior (Pickersgill, unpubl.; Eshbaugh, 1975). We use the technique of starch gel electrophoresis to provide data for a different perspective of evolution in Capsicum. Re-