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Julia Rueda

Other affiliations: University at Albany, SUNY
Bio: Julia Rueda is an academic researcher from Complutense University of Madrid. The author has contributed to research in topics: Gene & Gene expression. The author has an hindex of 13, co-authored 25 publications receiving 1307 citations. Previous affiliations of Julia Rueda include University at Albany, SUNY.

Papers
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Journal ArticleDOI
TL;DR: This article describes a network experiment involving several European laboratories, in which the reproducibility of three popular molecular marker techniques was examined: random-amplified fragment length polymorphism (RAPD), amplified fragment length SNP (AFLP) and sequence-tagged microsatellites (SSR).
Abstract: A number of PCR-based techniques can be used to detect polymorphisms in plants. For their wide-scale usage in germplasm characterisation and breeding it is important that these marker technologies can be exchanged between laboratories, which in turn requires that they can be standardised to yield reproducible results, so that direct collation and comparison of the data are possible. This article describes a network experiment involving several European laboratories, in which the reproducibility of three popular molecular marker techniques was examined: random-amplified fragment length polymorphism (RAPD), amplified fragment length polymorphism (AFLP) and sequence-tagged microsatellites (SSR). For each technique, an optimal system was chosen, which had been standardised and routinely used by one laboratory. This system (genetic screening package) was distributed to different participating laboratories in the network and the results obtained compared with those of the original sender. Different experiences were gained in this exchange experiment with the different techniques. RAPDs proved difficult to reproduce. For AFLPs, a single-band difference was observed in one track, whilst SSR alleles were amplified by all laboratories, but small differences in their sizing were obtained.

895 citations

Journal ArticleDOI
TL;DR: The results suggest that sequences necessary for expression in pollen are present in a region from −100 to −54, while other sequences which amplify that expression reside between −260 and −100 and the −260 to −100 region contains sequences similar to other protein-binding domains reported for plants.
Abstract: We have previously reported the isolation and characterization of a gene (Zm13) from Zea mays which shows a pollen-specific pattern of expression. Stably transformed tobacco plants containing a reporter gene linked to portions of the Zm13 5' flanking region show correct temporal and spatial expression of the gene. Here we present a more detailed analysis of the 5' regions responsible for expression in pollen by utilizing a transient expression system. Constructs containing the beta-glucuronidase (GUS) gene under the control of various sized fragments of the Zm 13 5' flanking region were introduced into Tradescantia and Zea mays pollen via high-velocity microprojectile bombardment, and monitored both visually and with a fluorescence assay. The results suggest that sequences necessary for expression in pollen are present in a region from -100 to -54, while other sequences which amplify that expression reside between -260 and -100. The replacement of the normal terminator with a portion of the Zm13 3' region containing the putative polyadenylation signal and site also increased GUS expression. While the -260 to -100 region contains sequences similar to other protein-binding domains reported for plants, the -100 to -54 region appears to contain no significant homology to other known promoter fragments which direct pollen-specific expression. The microprojectile bombardment of Tradescantia pollen appears to be a good test system for assaying maize and possibly other monocot promoter constructs for pollen expression.

107 citations

Book ChapterDOI
01 Jan 1998
TL;DR: A network reproducibility test was carried by nine European laboratories to determine whether RAPD profiles could be reproduced by different laboratories if all details of the reaction conditions were standardized.
Abstract: For the application of molecular techniques to wide-scale diversity screening it is important that the different screening techniques employed can be standardized to yield reproducible results across laboratories in order that direct collation and comparison of the data are possible (1). RAPDs (Random Amplified Polymorphic DNA) involve the use of a single ‘arbitrary’ primer (purchasable from commercial companies) in a PCR reaction and results in the amplification of several discrete DNA products (see Chapter 9). It is now widely recognized that to obtain reproducible band profiles on the gels it is absolutely essential to maintain consistent reaction conditions. Numerous studies have reported the separate effects of altering different parameters, such as ratio of template DNA/primers, concentration of Taq polymerase and Mg concentration, on the bands obtained (2–5). A network reproducibility test was carried by nine European laboratories (L1-L9) to determine whether RAPD profiles could be reproduced by different laboratories if all details of the reaction conditions were standardized (1).

40 citations

Journal ArticleDOI
TL;DR: Based on evidence from transient assays after microprojectile particle bombardment of the GmHSP 17·5E/GUS construct into pollen, it is likely that the gene is transcriptionally in an inactive configuration in pollen nuclei in stably transformed transgenic plants.
Abstract: A detailed histochemical analysis of the expression of the soybean small heat shock protein gene promoter, GmHSP 17·5E, fused to the GUS reporter gene, has been made in all organs and tissues of the flower as a function of stage of development and heat stress. This promoter is not uniformly expressed after a heat shock in all floral tissues and organs. Expression is seen at all stages of development in the sepals but not in the petals. The expression pattern in the pistil and in anthers is complex. Heat stress-induced GUS staining is seen in the style and upper portion of the ovary, but not in the stigmatic papillae or in the lower part of the ovary or in ovules. In stamens the heat shock response is seen in the filament and in the extension of the vascular tissue from the filament into the anther. No induction is seen in other tissues of the anther or in microspores or pollen at any stage of development. Vegetative organs in contrast are more uniform in the heat shock inducibility of GUS activity. Based on evidence from transient assays after microprojectile particle bombardment of the GmHSP 17·5E/GUS construct into pollen, it is likely that the gene is transcriptionally in an inactive configuration in pollen nuclei in stably transformed transgenic plants. These results are discussed with reference to other information in the literature.

39 citations

Book ChapterDOI
01 Jan 1998
TL;DR: Quantification of DNA is a very important step in many procedures where it is necessary to know the amount of DNA that is present when carrying out restriction digests or performing different techniques such as PCR and RAPDs.
Abstract: Quantification of DNA is a very important step in many procedures where it is necessary to know the amount of DNA that is present when carrying out restriction digests or performing different techniques such as PCR and RAPDs. There are several methods for quantifying DNA, the most widespread being: (i) the comparison of an aliquot of the extracted sample with standard DNAs of known concentration using gel electrophoresis and (ii) spectrophotometric determination. With both methods additional information is gained concerning the quality and purity of the extracted sample obtained. Normally both methods are used, but if the amount of DNA available is very small the gel electrophoresis technique alone may be employed. If there is no limitation on DNA amount, however, spectrophotometric measures should also be taken.

33 citations


Cited by
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Journal ArticleDOI
TL;DR: Four case studies representing a large variety of population genetics investigations differing in their sampling strategies, in the type of organism studied (plant or animal) and the molecular markers used [microsatellites or amplified fragment length polymorphisms (AFLPs), and the estimated genotyping error rate are considered.
Abstract: Genotyping errors occur when the genotype determined after molecular analysis does not correspond to the real genotype of the individual under consideration. Virtually every genetic data set includes some erroneous genotypes, but genotyping errors remain a taboo subject in population genetics, even though they might greatly bias the final conclusions, especially for studies based on individual identification. Here, we consider four case studies representing a large variety of population genetics investigations differing in their sampling strategies (noninvasive or traditional), in the type of organism studied (plant or animal) and the molecular markers used [microsatellites or amplified fragment length polymorphisms (AFLPs)]. In these data sets, the estimated genotyping error rate ranges from 0.8% for microsatellite loci from bear tissues to 2.6% for AFLP loci from dwarf birch leaves. Main sources of errors were allelic dropouts for microsatellites and differences in peak intensities for AFLPs, but in both cases human factors were non-negligible error generators. Therefore, tracking genotyping errors and identifying their causes are necessary to clean up the data sets and validate the final results according to the precision required. In addition, we propose the outline of a protocol designed to limit and quantify genotyping errors at each step of the genotyping process. In particular, we recommend (i) several efficient precautions to prevent contaminations and technical artefacts; (ii) systematic use of blind samples and automation; (iii) experience and rigor for laboratory work and scoring; and (iv) systematic reporting of the error rate in population genetics studies.

1,391 citations

Journal ArticleDOI
TL;DR: A protocol for estimating error rates is proposed and it is recommended that these measures be systemically reported to attest the reliability of published genotyping studies.
Abstract: Although genotyping errors affect most data and can markedly influence the biological conclusions of a study, they are too often neglected. Errors have various causes, but their occurrence and effect can be limited by considering these causes in the production and analysis of the data. Procedures that have been developed for dealing with errors in linkage studies, forensic analyses and non-invasive genotyping should be applied more broadly to any genetic study. We propose a protocol for estimating error rates and recommend that these measures be systemically reported to attest the reliability of published genotyping studies.

1,143 citations

Journal ArticleDOI
TL;DR: Because of their high replicability and ease of use, AFLP markers have emerged as a major new type of genetic marker with broad application in systematics, pathotyping, population genetics, DNA fingerprinting and quantitative trait loci (QTL) mapping.
Abstract: Amplified fragment length polymorphisms (AFLPs) are polymerase chain reaction (PCR)-based markers for the rapid screening of genetic diversity. AFLP methods rapidly generate hundreds of highly replicable markers from DNA of any organism; thus, they allow high-resolution genotyping of fingerprinting quality. The time and cost efficiency, replicability and resolution of AFLPs are superior or equal to those of other markers [allozymes, random amplified polymorphic DNA (RAPD), restriction fragment length polymorphism (RFLP), microsatellites], except that AFLP methods primarily generate dominant rather than co-dominant markers. Because of their high replicability and ease of use, AFLP markers have emerged as a major new type of genetic marker with broad application in systematics, pathotyping, population genetics, DNA fingerprinting and quantitative trait loci (QTL) mapping.

992 citations

Journal ArticleDOI
TL;DR: A synthesis of areas of AFLP technique, including comparison to other genotyping methods, assessment of errors, homoplasy, phylogenetic signal and appropriate analysis techniques are provided, with the aim of providing a review that will be applicable to all AFLP-based studies.

631 citations

Journal ArticleDOI
TL;DR: It is proposed that this method could be used in conjunction with RAPD markers for applications such as genetic analysis, bulked segregant analysis, and quantitative trait loci mapping, especially in laboratories with a preference for agarose gel electrophoresis.
Abstract: Random amplified polymorphic DNA (RAPD) markers have been used for numerous applications in plant molecular genetics research despite having disadvantages of poor reproducibility and not generally being associated with gene regions. A novel method for generating plant DNA markers was developed based on the short conserved region flanking the ATG start codon in plant genes. This method uses single 18-mer primers in single primer polymerase chain reaction (PCR) and an annealing temperature of 50°C. PCR amplicons are resolved using standard agarose gel electrophoresis. This method was validated in rice using a genetically diverse set of genotypes and a backcross population. Reproducibility was evaluated by using duplicate samples and conducting PCR on different days. Start codon targeted (SCoT) markers were generally reproducible but exceptions indicated that primer length and annealing temperature are not the sole factors determining reproducibility. SCoT marker PCR amplification profiles indicated dominant marker like RAPD markers. We propose that this method could be used in conjunction with these markers for applications such as genetic analysis, bulked segregant analysis, and quantitative trait loci mapping, especially in laboratories with a preference for agarose gel electrophoresis.

589 citations