Identification and Characterization of EI (Elongated Internode) Gene in Tomato (Solanum lycopersicum)

Internode length is an important agronomic trait affecting plant architecture and crop yield. However, few genes for internode elongation have been identified in tomato. In this study, we characterized an elongated internode inbred line P502, which is a natural mutant of the tomato cultivar 05T606. The mutant P502 exhibits longer internode and higher bioactive GA concentration compared with wild-type 05T606. Genetic analysis suggested that the elongated internode trait is controlled by quantitative trait loci (QTL). Then, we identified a major QTL on chromosome 2 based on molecular markers and bulked segregant analysis (BSA). The locus was designated as EI (Elongated Internode), which explained 73.6% genetic variance. The EI was further mapped to a 75.8-kb region containing 10 genes in the reference Heinz 1706 genome. One single nucleotide polymorphism (SNP) in the coding region of solyc02g080120.1 was identified, which encodes gibberellin 2-beta-dioxygenase 7 (SlGA2ox7). SlGA2ox7, orthologous to AtGA2ox7 and AtGA2ox8, is involved in the regulation of GA degradation. Overexpression of the wild EI gene in mutant P502 caused a dwarf phenotype with a shortened internode. The difference of EI expression levels was not significant in the P502 and wild-type, but the expression levels of GA biosynthetic genes including CPS, KO, KAO, GA20ox1, GA20ox2, GA20ox4, GA3ox1, GA2ox1, GA2ox2, GA2ox4, and GA2ox5, were upregulated in mutant P502. Our results may provide a better understanding of the genetics underlying the internode elongation and valuable information to improve plant architecture of the tomato.

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Biochemical and Structural Characterization of Amy1: An Alpha-Amylase from Cryptococcus flavus Expressed in Saccharomyces cerevisiae

Enzyme Research ◽

10.4061/2011/157294 ◽

2011 ◽

Vol 2011 ◽

pp. 1-7 ◽

Cited By ~ 14

Author(s):

Alexsandro Sobreira Galdino ◽

Roberto Nascimento Silva ◽

Muriele Taborda Lottermann ◽

Alice Cunha Morales Álvares ◽

Lídia Maria Pepe de Moraes ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Soluble Starch ◽

Alpha Amylase ◽

Enzyme Specificity ◽

Wild Type ◽

Spectra Analysis ◽

Exclusion Chromatography ◽

The Difference ◽

Thermal Stability Of

An extracellular alpha-amylase (Amy1) whose gene from Cryptococcus flavus was previously expressed in Saccharomyces cerevisiae was purified to homogeneity (67 kDa) by ion-exchange and molecular exclusion chromatography. The enzyme was activated by NH4+ and inhibited by Cu+2 and Hg+2. Significant biochemical and structural discrepancies between wild-type and recombinant α-amylase with respect to Km values, enzyme specificity, and secondary structure content were found. Far-UV CD spectra analysis at pH 7.0 revealed the high thermal stability of both proteins and the difference in folding pattern of Amy1 compared with wild-type amylase from C. flavus, which reflected in decrease (10-fold) of enzymatic activity of recombinant protein. Despite the differences, the highest activity of Amy1 towards soluble starch, amylopectin, and amylase, in contrast with the lowest activity of Amy1w, points to this protein as being of paramount biotechnological importance with many applications ranging from food industry to the production of biofuels.

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Absorption of cadmium accompanied by EDTA varies according to tomato cultivar

Crop and Pasture Science ◽

10.1071/cp19130 ◽

2019 ◽

Vol 70 (11) ◽

pp. 981

Author(s):

Shouping Zhao ◽

Qi Zhang ◽

Wendan Xiao ◽

De Chen ◽

Xuezhu Ye

Keyword(s):

Uptake Rate ◽

Plant Genotype ◽

Tomato Cultivar ◽

Cd Uptake ◽

Cd Accumulation ◽

Expression Levels ◽

Uptake Rates ◽

The Difference ◽

Cd Transport

Two tomato (Lycopersicum solanum) varieties, one high (YSL189) and one low (HZ903) cadmium (Cd) accumulator, were used in our experiment. We detected cadmium (Cd ion/Cd-EDTA) uptake rates in roots and the corresponding expression of the Cd transport genes IRT1, IRT2, ZIP, Nramp1, Nramp2 and Nramp3. Our data proved that both cultivars – YSL189 and HZ903 – showed higher Cd accumulation in plants and a higher Cd uptake rate in roots supplied with Cd ion than with Cd-EDTA. In roots of YSL189, the expression levels of IRT1, IRT2, ZIP, Nramp1, Nramp3 and Nramp2 (5, 10 and 20 µmolc L–1 Cd) treated with Cd ion were higher than those treated with Cd-EDTA, whereas in roots of HZ903, only two genes, IRT1 and Nramp1 (5, 10, 100 µmolc L–1 Cd), showed higher expression levels in plants treated with Cd ion than in those treated with Cd-EDTA. When the difference between the cultivars was considered, the Cd concentration in the plant and the Cd uptake rate in the roots of YSL189 were higher than those of YZ903 under the same Cd treatments (i.e. Cd ion or Cd-EDTA). The expression of IRT2 and ZIP in the roots of YSL189 was higher than that observed in HZ903 treated with all levels of ion-Cd. We attribute the higher Cd uptake rate and greater accumulation of ion-Cd compared with EDTA-Cd in YSL189 than those found in HZ903 partly to the genes that had higher expression levels. Our results indicate that the roles of transporters in the absorption of different forms of Cd vary according to plant genotype.

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Sequence Characterization of theMC1RGene in Yak (Poephagus grunniens) Breeds with Different Coat Colors

Journal of Biomedicine and Biotechnology ◽

10.1155/2009/861046 ◽

2009 ◽

Vol 2009 ◽

pp. 1-6 ◽

Cited By ~ 10

Author(s):

Shi-Yi Chen ◽

Yi Huang ◽

Qing Zhu ◽

Luca Fontanesi ◽

Yong-Gang Yao ◽

...

Keyword(s):

Coat Color ◽

Extracellular Loop ◽

Wild Type ◽

Coding Region ◽

Functional Effect ◽

Nucleotide Substitutions ◽

Melanocortin 1 Receptor ◽

Sequence Characterization ◽

Potential Association

Melanocortin 1 receptor (MC1R) gene plays a key role in determining coat color in several species, including the cattle. However, up to now there is no report regarding theMC1Rgene and the potential association of its mutations with coat colors in yak (Poephagus grunniens). In this study, we sequenced the encoding region of theMC1Rgene in three yak breeds with completely white (Tianzhu breed) or black coat color (Jiulong and Maiwa breeds). The predicted coding region of the yakMC1Rgene resulted of 954 bp, the same to that of the wild-type cattle sequence, with >99% identity. None of the mutation events reported in cattle was found. Comparing the yak obtained sequences, five nucleotide substitutions were detected, which defined three haplotypes (EY1,EY2, andEY3). Of the five mutations, two, characterizing theEY1haplotype, were nonsynonymous substitutions (c.340C>A and c.871G>A) causing amino acid changes located in the first extracellular loop (p.Q114K) and in the seventh transmembrane region (p.A291T).In silicoprediction might indicate a functional effect of the latter substitution. However, all three haplotypes were present in the three yak breeds with relatively consistent frequency distribution, despite of their distinguished coat colors, which suggested that there was no across-breed association between haplotypes or genotypes and black/white phenotypes, at least in the investigated breeds. Other genes may be involved in affecting coat color in the analyzed yaks.

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Postharvest Analysis of Lowland Transgenic Tomato Fruits Harboring hpRNAi-ACO1Construct

The Scientific World JOURNAL ◽

10.1100/2012/439870 ◽

2012 ◽

Vol 2012 ◽

pp. 1-9 ◽

Cited By ~ 8

Author(s):

Bita Behboodian ◽

Zainon Mohd Ali ◽

Ismanizan Ismail ◽

Zamri Zainal

Keyword(s):

Acc Oxidase ◽

Pectin Methylesterase ◽

Flower Senescence ◽

35S Promoter ◽

Specific Gene ◽

Wild Type ◽

Coding Region ◽

Tomato Cultivar ◽

Titratable Acid ◽

Rnai Technology

The plant hormone, ethylene, is an important regulator which involved in regulating fruit ripening and flower senescence. In this study, RNA interference (RNAi) technology was employed to silence the genes involved in ethylene biosynthetic pathway. This was achieved by blocking the expression of specific gene encoding the ACC oxidase. Initially, cDNA corresponding toACO1of lowland tomato cultivar (MT1), which has high identity withACO1ofSolanum lycopersicumin GenBank, was cloned through RT-PCR. Using a partial coding region ofACO1, one hpRNAi transformation vector was constructed and expressed ectopically under the 35S promoter. Results showed that transgenic lines harboring the hpRNA-ACO1construct had lower ethylene production and a longer shelf life of 32 days as compared to 10 days for wild-type fruits. Changes in cell wall degrading enzyme activities were also investigated in cases where the transgenic fruits exhibited reduced rates of firmness loss, which can be associated with a decrease in pectin methylesterase (PME) and polygalacturonase (PG) activities. However, no significant change was detected in both transgenic and wild-type fruits in terms ofβ-galactosidase (β-Gal) activity and levels of total soluble solid, titratable acid and ascorbic acid.

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Nucleotide variation and divergence in the histone multigene family in Drosophila melanogaster.

Genetics ◽

10.1093/genetics/122.1.87 ◽

1989 ◽

Vol 122 (1) ◽

pp. 87-97

Author(s):

Y Matsuo ◽

T Yamazaki

Keyword(s):

Drosophila Melanogaster ◽

Gene Family ◽

Purifying Selection ◽

Noncoding Region ◽

Chromosome 2 ◽

Coding Region ◽

Phylogenetic Relations ◽

Selection For ◽

The Difference ◽

Histone H3 Gene

Abstract Nucleotide differences in the histone H3 gene family in Drosophila melanogaster were studied on three levels: (1) within a chromosome, (2) within a population and (3) between species (D. melanogaster and Drosophila simulans). The average difference within the H3 gene within a chromosome was 0.0040 per nucleotide site, about 52% of that within a population (0.0077). The proportion of divergent sites between the two species was 0.0575, which is about 8.5 times the difference within a species. The distribution of divergence between species was similar to that of variation within a species. Divergence and variation were noted to be greatest in the 3' noncoding region and least in the coding region. Values intermediate between these were found for the 5' noncoding region. Divergence and variation in silent sites exceeded those in the total coding region, thus indicating possible purifying selection for amino-acid-altering change. Phylogenetic relations among H3 genes and genetic differences on these three levels are evidence for the concerted evolution of the histone gene family. The molecular mechanism by which variation is produced and maintained is discussed.

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Functional characterization of the chlorzoxazone 6-hydroxylation activity of human cytochrome P450 2E1 allelic variants in Han Chinese

PeerJ ◽

10.7717/peerj.9628 ◽

2020 ◽

Vol 8 ◽

pp. e9628

Author(s):

Ting Wang ◽

Huihui Du ◽

Jingsong Ma ◽

Lu Shen ◽

Muyun Wei ◽

...

Keyword(s):

Enzyme Activity ◽

Cytochrome P450 ◽

Genetic Polymorphisms ◽

Han Chinese ◽

Enzyme Function ◽

Wild Type ◽

Coding Region ◽

Allelic Variants ◽

Hydroxylation Activity

Backgrounds Cytochrome P450 (P450) 2E1 is one of the primary enzymes responsible for the metabolism of xenobiotics, such as drugs and environmental carcinogens. The genetic polymorphisms of the CYP2E1 gene in promoter and coding regions have been identified previously in the Han Chinese population from four different geographic areas of Mainland China. Methods To investigate whether genetic variants identified in the CYP2E1 coding region affect enzyme function, the enzymes of four single nucleotide polymorphism (SNP) variants in the coding region (novel c.1009C>T, causing p.Arg337X, where X represents the translational stop codon; c.227G>A, causing p.Arg76His; c.517G>A, yielding p.Gly173Ser; and c.1263C>T, presenting the highest allele frequency), two novel alleles (c.[227G>A;1263C>T] and c.[517G>A;1263C>T]), and the wild-type CYP2E1 were heterologously expressed in COS-7 cells and functionally characterized in terms of expression level and chlorzoxazone 6-hydroxylation activity. The impact of the CYP2E1 variant sequence on enzyme activity was predicted with three programs: Polyphen 2, PROVEAN and SIFT. Results The prematurely terminated p.Arg337X variant enzyme was undetectable by western blotting and inactive toward chlorzoxazone 6-hydroxylation. The c.1263C>T and c.[517G>A;1263C>T] variant enzymes exhibited properties similar to those of the wild-type CYP2E1. The CYP2E1 variants c.227G>A and c.[227G>A;1263C>T] displayed significantly reduced enzyme activity relative to that of the wild-type enzyme (decreased by 42.8% and 32.8%, respectively; P < 0.01). The chlorzoxazone 6-hydroxylation activity of the c.517G>A transfectant was increased by 31% compared with the wild-type CYP2E1 enzyme (P < 0.01). Positive correlations were observed between the protein content and enzyme activity for CYP2E1 (P = 0.0005, r2 = 0.8833). The characterization of enzyme function allelic variants in vitro was consistent with the potentially deleterious effect of the amino acid changes as determined by prediction tools. Conclusions These findings indicate that the genetic polymorphisms of CYP2E1, i.e., c.1009C>T (p.Arg337X), c.227G>A (p.Arg76His), and c.517G>A (p.Gly173Ser), could influence the metabolism of CYP2E1 substrates, such as chlorzoxazone.

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Characterization of an Isogenic Mutant ofStreptococcus pyogenes Manfredo Lacking the Ability To Make Streptococcal Acid Glycoprotein

Infection and Immunity ◽

10.1128/iai.68.5.2441-2448.2000 ◽

2000 ◽

Vol 68 (5) ◽

pp. 2441-2448 ◽

Cited By ~ 89

Author(s):

B. A. Degnan ◽

M. C. Fontaine ◽

A. H. Doebereiner ◽

J. J. Lee ◽

P. Mastroeni ◽

...

Keyword(s):

Human Peripheral Blood ◽

Arginine Deiminase ◽

Significant Activity ◽

Wild Type ◽

Peripheral Blood Mononuclear ◽

Isogenic Mutant ◽

Acid Glycoprotein ◽

Cell Extracts ◽

The Difference

ABSTRACT An isogenic mutant of Streptococcus pyogenes Manfredo that lacks the ability to make streptococcal acid glycoprotein (SAGP) has been constructed by inserting a deletion in the sagpgene using the method of allelic exchange. An assay of cell extracts (CE) prepared from the wild-type and mutant Manfredo strains for the enzyme arginine deiminase (AD) showed that significant activity was present in wild-type CE but none could be detected in mutant CE. These findings confirm our earlier conclusion that SAGP has AD activity (B. A. Degnan, J. M. Palmer, T. Robson, C. E. D. Jones, M. Fischer, M. Glanville, G. D. Mellor, A. G. Diamond, M. A. Kehoe, and J. A. Goodacre, Infect. Immun. 66:3050–3058, 1998). Wild-type CE but not mutant CE potently inhibited human peripheral blood mononuclear cell proliferation in response to phytohemagglutinin, and this inhibition was overcome by the addition ofl-arginine to proliferation assay mixtures. Invasion assays showed that the isogenic mutant organisms lacking SAGP, and thus AD activity, were between three and five times less able to enter epithelial cells (Hep-2C and A549) than were the wild-type streptococci. Both wild-type and mutant S. pyogenesbacteria were extremely sensitive to low pH. However,l-arginine (1 mM or above) significantly increased the viability of the wild type but not the isogenic mutant organisms under acidic conditions. The difference in acid susceptibility between wild-type and mutant bacteria may explain the reduced capacity of the isogenic mutant bacteria to invade and survive intracellularly.

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Characterization of a Listeria monocytogenes Scott A Isolate with High Tolerance towards High Hydrostatic Pressure

Applied and Environmental Microbiology ◽

10.1128/aem.68.7.3183-3189.2002 ◽

2002 ◽

Vol 68 (7) ◽

pp. 3183-3189 ◽

Cited By ~ 85

Author(s):

Kimon A. G. Karatzas ◽

Marjon H. J. Bennik

Keyword(s):

Hydrostatic Pressure ◽

High Hydrostatic Pressure ◽

Exponential Phase ◽

Wild Type ◽

Food Borne ◽

Lower Maximum ◽

The Difference ◽

Specific Growth Rates

ABSTRACT An isolate of L. monocytogenes Scott A that is tolerant to high hydrostatic pressure (HHP), named AK01, was isolated upon a single pressurization treatment of 400 MPa for 20 min and was further characterized. The survival of exponential- and stationary-phase cells of AK01 in ACES [N-(2-acetamido)-2-aminoethanesulfonic acid] buffer was at least 2 log units higher than that of the wild type over a broad range of pressures (150 to 500 MPa), while both strains showed higher HHP tolerance (piezotolerance) in the stationary than in the exponential phase of growth. In semiskim milk, exponential-phase cells of both strains showed lower reductions upon pressurization than in buffer, but again, AK01 was more piezotolerant than the wild type. The piezotolerance of AK01 was retained for at least 40 generations in rich medium, suggesting a stable phenotype. Interestingly, cells of AK01 lacked flagella, were elongated, and showed slightly lower maximum specific growth rates than the wild type at 8, 22, and 30°C. Moreover, the piezotolerant strain AK01 showed increased resistance to heat, acid, and H2O2 compared with the wild type. The difference in HHP tolerance between the piezotolerant strain and the wild-type strain could not be attributed to differences in membrane fluidity, since strain AK01 and the wild type had identical in situ lipid melting curves as determined by Fourier transform infrared spectroscopy. The demonstrated occurrence of a piezotolerant isolate of L. monocytogenes underscores the need to further investigate the mechanisms underlying HHP resistance of food-borne microorganisms, which in turn will contribute to the appropriate design of safe, accurate, and feasible HHP treatments.

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Characterization of the Original Christmas Disease Mutation (Cysteine 206 → Serine): From Clinical Recognition to Molecular Pathogenesis

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648381 ◽

1992 ◽

Vol 67 (01) ◽

pp. 063-065 ◽

Cited By ~ 6

Author(s):

Sherryl A M Taylor ◽

Jacalyn Duffin ◽

Cherie Cameron ◽

Jerome Teitel ◽

Bernadette Garvey ◽

...

Keyword(s):

Nucleotide Substitution ◽

Novel Mutation ◽

Factor Ix ◽

Causative Mutation ◽

Coding Region ◽

Body Of Knowledge ◽

Polymerase Chain ◽

Splice Junctions

SummaryChristmas disease was first reported as a distinct clinical entity in two manuscripts published in 1952 (1, 2). The eponym associated with this disorder, is the surname of the first patient examined in detail and reported by Biggs and colleagues in a paper describing the clinical and laboratory features of seven affected individuals (3). This patient has severe factor IX coagulant deficiency (less than 0.01 units/ml) and no detectable circulating factor IX antigen (less than 0.01 units/ml). Coding sequence and splice junctions of the factor IX gene from this patient have been amplified in vitro through the polymerase chain reaction (PCR). One nucleotide substitution was identified at nucleotide 30,070 where a guanine was replaced by a cytosine. This mutation alters the amino acid encoded at position 206 in the factor IX protein from cysteine to serine. The non conservative nature of this substitution, the absence of this change in more than 200 previously sequenced factor IX genes and the fact that the remainder of the coding region of this gene was normal, all provide strong circumstantial evidence in favour of this change being the causative mutation in this patient. The molecular characterization of this novel mutation in the index case of Christmas disease, contributes to the rapidly expanding body of knowledge pertaining to Christmas disease pathogenesis.

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