Autocrine stimulation after transfer of the granulocyte/macrophage colony-stimulating factor gene and autonomous growth are distinct but interdependent steps in the oncogenic pathway ( ABSTRACTAutocrine stimulation of cells by aberrant synthesis of growth factor may lead to malignant transformation, either as a direct consequence of endogenous factor production or as a first step of a series of successive events.

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We show that a 2.6 kb fragment of the muscle myosin heavy-chain gene (Mhc) of Drosophila melanogaster (containing 458 base pairs of upstream sequence, the first exon, the first intron and the beginning of the second exon) drives expression in all muscles. Comparison of the minimal promoter to Mhc genes of ten Drosophila species identified putative regulatory elements in the upstream region and in the first intron. The first intron is required for expression in four small cells of the tergal depressor of the trochanter (jump) muscle and in the indirect flight muscle. The 3′ end of this intron is important for Mhc transcription in embryonic body wall muscle and contains AT-rich elements that are protected from DNase I digestion by nuclear proteins of Drosophila embryos. Sequences responsible for expression in embryonic, adult body wall and adult head muscles are present both within and outside the intron. Elements important for expression in leg muscles and in the large cells of the jump muscle flank the intron. We conclude that multiple transcriptional regulatory elements are responsible for Mhc expression in specific sets of Drosophila muscles. KeywordsDrosophila melanogaster; muscle; myosin heavy chain; transcription; enhancer; gene regulation Drosophila melanogaster is unusual in having a single muscle myosin heavy-chain gene (Mhc) (Bernstein et al., 1983;Rozek and Davidson, 1983), rather than a Mhc multigene family (see Emerson and Bernstein, 1987 for review). Alternative splicing of the primary transcripts from this gene yields multiple isoforms of the protein (Collier et al., 1990;George et al., 1989;Hastings and Emerson, 1991;Kazzaz and Rozek, 1989;Kronert et al., 1991;Zhang and Bernstein, 2001). Drosophila Mhc is expressed in all muscles at embryonic, larval, pupal and adult stages. Thus several stage-or muscle-specific enhancer elements may regulate its transcription.Transcriptional regulatory regions for a number of Drosophila muscle genes have been mapped, resulting in the identification of both unique and shared cis-acting elements (Arredondo et al., 2001;Kelly et al., 2002;Marin et al., 2004;Mas et al., 2004;Meredith and Storti, 1993). The latter includes E-boxes that bind helix-loop-helix transcription factors and MEF2 sites that bind MEF2 protein (Bour et al., 1995; Lilly et al., 1995). E-boxes, MEF2 sites and their binding factors positively regulate vertebrate muscle gene expression as well (see Molkentin and Olson, 1996 for review).* Corresponding author. Tel.: +1-619-594-5629; fax: +1-619-594-5676; E-mail address: sbernst@sunstroke.sdsu.edu. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal dis...

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A new screening method for the detection and identification of GMO, based on the use of multiplex PCR followed by microarray, has been developed and is presented. The technology is based on the identification of quite ubiquitous GMO genetic target elements first amplified by PCR, followed by direct hybridisation of the amplicons on a predefined microarray (DualChip Ò GMO, Eppendorf, Germany). The validation was performed within the framework of a European project (Co-Extra, contract no 007158) and in collaboration with 12 laboratories specialised in GMO detection. The present study reports the strategy and the results of an ISO complying validation of the method carried out through an inter-laboratory study. Sets of blind samples were provided consisting of DNA reference materials covering all the elements detectable by specific probes present on the array. The GMO concentrations varied from 1% down to 0.045%. In addition, a mixture of two GMO events (0.1% RRS diluted in 100% TOPAS19/2) was incorporated in the S. Leimanis Á S. Hamels (&) Á J. Remacle Eppendorf Array Technologies, 20 Rue du séminaire, 5000 Namur, Belgium e-mail: hamels.s@eppendorf.be DOI 10.1007/s00217-008-0886-y study to test the robustness of the assay in extreme conditions. Data were processed according to ISO 5725 standard. The method was evaluated with predefined performance criteria with respect to the EC CRL method acceptance criteria. The overall method performance met the acceptance criteria; in particular, the results showed that the method is suitable for the detection of the different target elements at 0.1% concentration of GMO with a 95% accuracy rate. This collaborative trial showed that the method can be considered as fit for the purpose of screening with respect to its intra-and inter-laboratory accuracy. The results demonstrated the validity of combining multiplex PCR with array detection as provided by the DualChip Ò GMO (Eppendorf, Germany) for the screening of GMO. The results showed that the technology is robust, practical and suitable as a screening tool.123 Eur Food Res Technol

Monitoring of market products for detection of genetically modified organisms (GMO) is needed to comply with legislation in force in many regions of the world, to enforce traceability and to allow official control along the production and the distribution chains. This objective can be more easily achieved if reliable, time and cost-effective analytical methods are available. A GMO can be detected using either DNA-based or protein-based methods; both present advantages and disadvantages. The objective of this work was to assess the performance of a protein-based (lateral flow strips-LFT) and of a DNA-based (polymerase chain reaction-PCR) detection method for GMO analysis. One thousand five hundred samples of soybean, deriving from the sampling of 15 independent bulk lots in large M. Mazzara (*) : E. Grazioli : S. Larcher : G. Van -012-9407-5 shipments, were analysed to assess and compare the performance of the analytical methods and evaluate their suitability for GMO testing. Several indicators were used to compare the performance of the methods, including the percentage correlation between the PCR and LFT results. The GMO content of the samples ranged from 0 up to 100 %, allowing a full assessment of both analytical approaches with respect to all possible GMO content scenarios. The study revealed a very similar performance of the two methodologies, with low false-negative and false-positive results, and a very satisfactory capacity of both methods in detecting low amounts of target. While determining the fitness for purpose of both analytical approaches, this study also underlines the importance of alternative method characteristics, like costs and time.Anal. Methods (2013) 6:210-220 DOI 10.1007/s12161