Length, orientation, and plant host influence the mutation frequency in microsatellites

Microsatellites are simple, tandem DNA repeats that represent unstable regions of the genome. They undergo frequent changes in tract length by base additions or deletions due to DNA polymerase slippage during replication. To characterize factors affecting the frequency of spontaneous mutations occurring in microsatellites in plants, a reporter system was used in Arabidopsis thaliana and tomato ( Lycopersicon esculentum ). The β-glucuronidase (GUS) reporter system was used to measure the mutation frequency in various microsatellites (G7, G10, G13, G16, and C16) in somatic tissues. Our results indicate that this frequency increases with the number of repeats: a G16 tract was almost 80-fold more mutable than a G7 tract. Furthermore, the frequency of mutations depends on repeat orientation, as G16 was 3-fold more mutable than C16. The mutation rate was also found to differ markedly in Arabidopsis and tomato for an identical microsatellite. Indeed, Arabidopsis showed a 5-fold higher mutation frequency than tomato with the same G7 reporter construct. Finally, mutation in a G16 tract was frequent enough that mutations transmitted germinally to the next generation could be detected at a relatively high frequency.

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Bioactivity guided isolation of phytoestrogenic compounds from Cyclopia genistoides by the pER8:GUS reporter system

South African Journal of Botany ◽

10.1016/j.sajb.2016.06.001 ◽

2017 ◽

Vol 110 ◽

pp. 201-207 ◽

Cited By ~ 4

Author(s):

O. Roza ◽

W.-C. Lai ◽

I. Zupkó ◽

J. Hohmann ◽

N. Jedlinszki ◽

...

Keyword(s):

Reporter System ◽

Gus Reporter ◽

Gus Reporter System

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Mixed Inocula of N2-Fixing Bacteria on Summer Cereals: Utilisation of the gus Reporter System to Follow the Bacterial Plant Colonisation

Biological Nitrogen Fixation for the 21st Century - Current Plant Science and Biotechnology in Agriculture ◽

10.1007/978-94-011-5159-7_260 ◽

1998 ◽

pp. 415-415

Author(s):

D. Sticchi ◽

A. L. Javicoli ◽

C. Piccone ◽

P. Fabbri ◽

S. Mannelli ◽

...

Keyword(s):

Reporter System ◽

Plant Colonisation ◽

Gus Reporter ◽

Mixed Inocula ◽

Gus Reporter System

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The GUS Reporter System in Flower Development Studies

Methods in Molecular Biology - Flower Development ◽

10.1007/978-1-4614-9408-9_15 ◽

2013 ◽

pp. 295-304 ◽

Cited By ~ 6

Author(s):

Janaki S. Mudunkothge ◽

Beth A. Krizek

Keyword(s):

Flower Development ◽

Development Studies ◽

Reporter System ◽

Gus Reporter ◽

Gus Reporter System

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Satellite DNA in Plants: More than Just Rubbish

Cytogenetic and Genome Research ◽

10.1159/000437008 ◽

2015 ◽

Vol 146 (2) ◽

pp. 153-170 ◽

Cited By ~ 64

Author(s):

Manuel A. Garrido-Ramos

Keyword(s):

Dna Sequences ◽

Satellite Dna ◽

Dna Repeats ◽

Satellite Dnas ◽

Factors Affecting ◽

Functional Roles ◽

Tandem Repeat Sequences ◽

Satellite Repeats ◽

History Of ◽

Main Components

For decades, satellite DNAs have been the hidden part of genomes. Initially considered as junk DNA, there is currently an increasing appreciation of the functional significance of satellite DNA repeats and of their sequences. Satellite DNA families accumulate in the heterochromatin in different parts of the eukaryotic chromosomes, mainly in pericentromeric and subtelomeric regions, but they also span the functional centromere. Tandem repeat sequences may spread from subtelomeric to interstitial loci, leading to the formation of chromosome-specific loci or to the accumulation in equilocal sites in different chromosomes. They also appear as the main components of the heterochromatin in the sex-specific region of sex chromosomes. Satellite DNA, required for chromosome organization, also plays a role in pairing and segregation. Some satellite repeats are transcribed and can participate in the formation and maintenance of heterochromatin structure and in the modulation of gene expression. In addition to the identification of the different satellite DNA families, their characteristics and location, we are interested in determining their impact on the genomes, by identifying the mechanisms leading to their appearance and amplification as well as in understanding how they change over time, the factors affecting these changes, and the influence exerted by the evolutionary history of the organisms. On the other hand, satellite DNA sequences are rapidly evolving sequences that may cause reproductive barriers between organisms and promote speciation. The accumulation of experimental data collected in recent years and the emergence of new approaches based on next-generation sequencing and high-throughput genome analysis are opening new perspectives that are changing our understanding of satellite DNA. This review examines recent data to provide a timely update on the overall information gathered about this part of the genome, focusing on the advances in the knowledge of its origin, its evolution, and its potential functional roles.

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Chromosomal Location Plays a Role in Regulation of Aflatoxin Gene Expression in Aspergillus parasiticus

Applied and Environmental Microbiology ◽

10.1128/aem.68.1.306-315.2002 ◽

2002 ◽

Vol 68 (1) ◽

pp. 306-315 ◽

Cited By ~ 61

Author(s):

Ching-Hsun Chiou ◽

Michael Miller ◽

David L. Wilson ◽

Frances Trail ◽

John E. Linz

Keyword(s):

Gene Cluster ◽

Promoter Activity ◽

Aspergillus Parasiticus ◽

Chromosomal Location ◽

Selectable Marker ◽

Growth Conditions ◽

Gus Expression ◽

Reporter System ◽

Gus Reporter ◽

Homologous Site

ABSTRACT The nor-1 gene in the filamentous fungus Aspergillus parasiticus encodes a ketoreductase involved in aflatoxin biosynthesis. To study environmental influences on nor-1 expression, we generated plasmid pAPGUSNNB containing a nor-1 promoter-β-glucuronidase (GUS) (encoded by uidA) reporter fusion with niaD (encodes nitrate reductase) as a selectable marker. niaD transformants of A. parasiticus strain NR-1 (niaD) carried pAPGUSNNB integrated predominantly at the nor-1 or niaD locus. Expression of the native nor-1 and nor-1::GUS reporter was compared in transformants grown under aflatoxin-inducing conditions by Northern and Western analyses and by qualitative and quantitative GUS activity assays. The timing and level of nor-1 promoter function with pAPGUSNNB integrated at nor-1 was similar to that observed for the native nor-1 gene. In contrast, nor-1 promoter activity in pAPGUSNNB and a second nor-1::GUS reporter construct, pBNG3.0, was not detectable when integration occurred at niaD. Because niaD-dependent regulation could account for the absence of expression at niaD, a third chromosomal location was analyzed using pAPGUSNP, which contained nor-1::GUS plus pyrG (encodes OMP decarboxylase) as a selectable marker. GUS expression was detectable only when pAPGUSNP integrated at nor-1 and was not detectable at pyrG, even under growth conditions that required pyrG expression. nor-1::GUS is regulated similarly to the native nor-1 gene when it is integrated at its homologous site within the aflatoxin gene cluster but is not expressed at native nor-1 levels at two locations outside of the aflatoxin gene cluster. We conclude that the GUS reporter system can be used effectively to measure nor-1 promoter activity and that nor-1 is subject to position-dependent regulation in the A. parasiticus chromosome.

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Factors Affecting Agrobacterium-mediated Transformation of Common Bean

Journal of the American Society for Horticultural Science ◽

10.21273/jashs.122.3.300 ◽

1997 ◽

Vol 122 (3) ◽

pp. 300-305 ◽

Cited By ~ 18

Author(s):

Zhanyuan Zhang ◽

Dermot P. Coyne ◽

Amitava Mitra

Keyword(s):

Common Bean ◽

Filter Paper ◽

Blue Staining ◽

Transformation Rate ◽

Gus Gene ◽

Reporter System ◽

Factors Affecting ◽

Host Interactions ◽

Optimized Protocol ◽

High Concentrations

Factors influencing Agrobacterium tumefaciens-mediated transformation of common beans (Phaseolus vulgaris L.) were examined using an intron-containing β-glucuronidase (GUS) gene as a reporter system to develop a repeatable transformation protocol. Tissue culture procedures used were based on direct shoot organogenesis. Two A. tumefaciens strains—A2760 and EHA105—were used, with emphasis on the former due to its overall higher infection rate. Eleven common-bean genotypes were compared for susceptibility to strain A2760 or EHA105. The pinto bean `Othello' was used extensively in testing different transformation conditions. Factors significantly affecting transformation rate were Agrobacterium × host interactions, explant maturity, preculture and cocultivation conditions, and selection schemes, based on transient GUS gene expression. The best transformation conditions were the use of susceptible genotypes and explants derived from mature seeds, preconditioning of explants in a medium containing 20 μmol of benzyladenine (BA) in darkness or on a filter paper, dipping explants in high concentrations of Agrobacterium cell suspension (OD650 = 0.8-1.0) followed by a long-term (6-day) cocultivation period on a semisolid agar medium in the presence of cytokinin or 3-day cocultivation on a moistened filter paper, and the use of lethal levels of selective agents. About 4% of explants, or 14% of regenerated shoots or buds, were putatively transgenic, as indicated by GUS blue staining throughout the entire shoot or bud, after explants were transformed with Agrobacterium strain A2760 using an optimized protocol.

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Factors Affecting Agrobacterium-mediated Transformation of Common Bean (Phaseolus vulgaris L.)

HortScience ◽

10.21273/hortsci.31.4.616f ◽

1996 ◽

Vol 31 (4) ◽

pp. 616f-616

Author(s):

Zhanyuan Zhang ◽

A. Mitra ◽

D.P. Coyne

Keyword(s):

Shoot Organogenesis ◽

Transformation Rate ◽

Bacterial Strains ◽

Phaseolus Vulgaris L ◽

Reporter System ◽

Breeding Lines ◽

Factors Affecting ◽

Host Interactions ◽

High Selection ◽

Direct Shoot Organogenesis

Factors influencing Agrobacterium–mediated DNA transfer of P. vulgaris were examined using an intron-containing β-glucuronidase (GUS) gene as a reporter system. Tissue culture procedures used were based on direct shoot organogenesis. Two A. tumefaciens strains, A2760 and EHA105, were used with more emphasis on the former due to its overall higher transformation rate. Ten bean entries including breeding lines and cultivars from both Meso-American and Andean origins were compared for compatibility with the two bacterial strains under different pre- and coculture conditions. Pinto `Othello' was extensively used in testing different transformation conditions. Factors found to have significant effects on transformation rate included Agrobacterium-host interactions, explant maturity, preculture and cocultivation conditions, as well as selection schemes, based on transient expression. Some factors, such as the effect of explant maturity and dark preconditioning of explants on gene transfer, have not been reported before. The best transformation conditions included the use of susceptible genotypes and mature explants, preconditioning of explants in darkness, followed by a maximum cocultivation period in the presence of cytokinin, and the use of high selection pressure.

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The GUS Reporter System as a Tool to Study Plant Gene Expression

Gus Protocols ◽

10.1016/b978-0-12-274010-7.50008-2 ◽

1992 ◽

pp. 23-43 ◽

Cited By ~ 55

Author(s):

Thomas Martin ◽

Rosa-Valentina Wöhner ◽

Sabine Hummel ◽

Lothar Willmitzer ◽

Wolf B. Frommer

Keyword(s):

Gene Expression ◽

Reporter System ◽

Plant Gene Expression ◽

Plant Gene ◽

Gus Reporter ◽

Gus Reporter System

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Factors Affecting Inverted Repeat Stimulation of Recombination and Deletion in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/148.4.1507 ◽

1998 ◽

Vol 148 (4) ◽

pp. 1507-1524 ◽

Cited By ~ 9

Author(s):

Kirill S Lobachev ◽

Boris M Shor ◽

Hiep T Tran ◽

Wendy Taylor ◽

J Dianne Keen ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Secondary Structure ◽

Genetic Instability ◽

Inverted Repeats ◽

Dna Repeats ◽

Origin Of Replication ◽

Adjacent Region ◽

Factors Affecting ◽

Stimulation Of

AbstractInverted DNA repeats are an at-risk motif for genetic instability that can induce both deletions and recombination in yeast. We investigated the role of the length of inverted repeats and size of the DNA separating the repeats for deletion and recombination. Stimulation of both deletion and recombination was directly related to the size of inverted repeats and inversely related to the size of intervening spacers. A perfect palindrome, formed by two 1.0-kb URA3-inverted repeats, increased intra- and interchromosomal recombination in the adjacent region 2,400-fold and 17,000-fold, respectively. The presence of a strong origin of replication in the spacer reduced both rates of deletion and recombination. These results support a model in which the stimulation of deletion and recombination by inverted repeats is initiated by a secondary structure formed between single-stranded DNA of inverted repeats during replication.

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