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1

Application of bioinformatics in GMO detection

100%
Nenov A., Vassilev D.
Biotechnologia
|
2006
|
issue 3
24-35
EN
Intensified production and release of GMOs into the environment and market are correlated with the development of evaluation requirement and tools for GMO screening. In the last years, application of integrated approaches based on molecular detection techniques and bioinformatics tools has been noted. The examples of bioinformatics system, methods, and IT achievements are described in this paper.
2

Technology Luminex xMAP? ? a new tool in the diagnostics of plant diseases

80%
Sledz W., Los E., Lojkowska E.
Biotechnologia
|
2010
|
issue 1
97-108
EN
The presented paper reviews the aspects of Luminex xMAP? technology as a new tool in diagnostics of plant diseases. Luminex xMAP? technology, based on flow cytometry, employs tiny colored beads (microspheres) combined with specific analytes and ligands (antibodies, oligonulectydies ect.). Initially, this system was known as FlowMetrix. The Luminex xMAP? technology is mainly employed in medical diagnostics. This technology is suited to very wide range of medical applications: allergy testing, cancer markers, gene expression, genotyping, tissue typing and many others. Luminex xMAP? technology was applied in the diagnostics of plant diseases. The first attempts were directed towards the fungal and bacterial plant pathogens. Nowadays, this technology has been applied to detection of bacteria such as: Pectobacterium carotovorum or Dickeya dianthicola and viral pathogens such as: Potato Virus X, Potato Virus Y and Potato Leafroll Virus. The Luminex xMAP? technology offers simultaneous detection of several factors (pathogens) in one test.
3

A quantitative bacterial micro-assay for rapid detection of serum phenylalanine in dry blood-spots: Application in phenylketonuria screening

80%
Vallian S., Moeini H.
Journal of Applied Genetics
|
2006
|
vol. 47
|
issue 1
79-83
EN
Phenylketonuria is an inherited metabolic disease, which is characterized by increased level of serum phenylalanine (Phe). The quantitative measurement of Phe in the serum is necessary to confirm the disease, and to distinguish phenylketonuria from other forms of hyperphenylalaninemia. In this study, we report a rapid and inexpensive micro-assay for simultaneous detection and quantitative measurement of serum Phe in dry blood-spots. Analysis of the standard curve showed a broad linear Phe range of 120?1800 mumol L?1. Application of this method in conjunction with the standard Guthrie bacterial inhibition assay and high-pressure liquid chromatography in analyzing 34 samples from phenylketonuria patients and control samples produced comparable results, with the regression equation of Y= 0.994X + 0.996. The advantage of this method over the Guthrie bacterial inhibition assay is its ability to measure the serum Phe quantitatively without false positive results. The method was successfully applied to dried blood-spots as well as serum and whole blood samples. The cost per sample is about 20?50 US cents, which is much less than those of high-pressure liquid chromatography and enzymatic commercial kits. The method can be automated, which is suitable for neonatal and mass phenylketonuria screening, especially in developing countries, where funding is a limiting factor.
4

PCR in real time. The methods of data analysis

80%
Tyburski J., Studzinska A., Daca P., Tretyn A.
Biotechnologia
|
2008
|
issue 1
86-96
EN
The accuracy of real-time PCR (RT-PCR) experiment is strongly dependent on the mathematical models of data analysis, on which the quantitative methods are based. In this review, we discuss the key steps of analysing data from real-time PCR experiments. These are the treshold cycle determination, estimation of real-time PCR amplification efficiency and amplicon quantification. The fit point method and the second derivative method are commonly used to determine a treshold cycle value, which is a cycle number in the early exponential phase of PCR that is used to calculate the initial amount of template DNA. The amplification efficiency calculation is usually based on the data collected from a standard curve. However, in the alternative methods, the amplification efficiency of an individual reaction is calculated from the kinetics of the reaction. Quantification of amplicon levels can be either absolute or relative. In absolute quantification method, the initial concentration of target template in unknowns, based on their cycle treshold values, and the construction of an absolute standard curve for each individual amplicon is required. Relative quantification can be done by the use of the relative standard curve method or the comparative method. In the first one, the initial amount of unknown samples is calculated from the standard curve of specific gene and normalized to the input amount of a reference gene which is also calculated from the standard curve. The comparative method is a mathematical model that is based on the differences in the normalized amplicon levels between the unknowns and control sample. There are also models that combine gene quantification and normalization into a single calculation.
5

Molecular analysis in detection and identification of genetically modified organisms (GMO)

80%
Linkiewicz A., Wisniewska I., Sowa S.
Biotechnologia
|
2006
|
issue 3
44-52
EN
Several DNA and protein-based analytical methods for GMOs detection have been established so far. For the detection of GMOs at the DNA level, mostly PCR-based methods are used, whereas at protein level ELISA and lateral strips are predominant. The choice of target sequence motif is the most important factor controlling the specificity of the PCR method. This review summarizes most widely used GMO analysis technologies and point out new areas of analytical investigations.
6

Detection of Fusarium tricinctum from cereal grain using PCR assay

80%
Kulik T.
Journal of Applied Genetics
|
2008
|
vol. 49
|
issue 3
305-311
EN
Contamination of cereals with mycotoxins produced by Fusarium is a worldwide problem requiring rapid and sensitive detection methods. This paper describes the development of a PCR protocol facilitating the detection of F. tricinctum, which belongs to the FHB (Fusarium Head Blight) complex responsible for contamination of cereal grains with enniatins and moniliformin. Sequence alignment of partial IGS rDNA revealed a single nucleotide polymorphism, which was used to design primers differentiating F. tricinctum from other members of the FHB complex. The specificity of the assay was tested on 68 isolates belonging to 21 Fusarium species originating from different parts of the world and hosts/substrates. Positive PCR results were obtained from all 12 F. tricinctum isolates tested; however, unexpected amplicons were amplified from the templates of F. acuminatum (CBS 618.87) and F. nurragi (CBS 393.96). No cross reactivity was found with any other Fusarium species tested. The PCR assay was tested on 24 asymptomatic wheat seed samples originating from Northern Poland and resulted in 13 positive samples, of which 11 samples were contaminated with moniliformin and/or antibiotic Y.
7

Apoptosis in mammalian sperm

70%
Trzcinska M.
Biotechnologia
|
2006
|
issue 1
82-89
EN
This physiological cell death, also known as programmed cell death, takes place during the entire growth period of an organism. In general, somatic cell apoptosis can be induced through extrinsic mechanisms acting at the plasma membrane, mitochondrial or nuclear levels. Recent studies have demonstrated that apoptosis is an underlying mechanism of germ cell death during normal spermatogenesis and that it is a major mechanism in regulating spermatogenesis of various mammalian species. While apoptosis in somatic cells and in (testicular) spermatocytes and spermatids is well established, the presence and significance of apoptosis in ejaculated animals sperm is still unresolved.
8

The control and eradication of bacterial infections and contaminations in plant tissue culture

70%
Orlikowska T., Sobiczewski P., Zawadzka M., Zenkteler E.
Biotechnologia
|
2010
|
issue 2
57-71
EN
Bacterial contamination is a serious problem in plant tissue culture. In in vitro cultures, if bacteria are introduced, it is most frequently with the initial explants, but bacterial contamination can also come from the laboratory environment or from the staff themselves. Exogenous bacteria are easier to deal with, but endogenous bacteria remain problematic. Standard sterilization with ethanol, NaOCl or HgCl2 and with antibiotics can now be enriched with new components (NaDCC, AgNO3, nano-silver) or sanitation products (PPMTM, ProClin? 300, Biosept 33 SL, Vitrofural?, Dekaben). Some of these can be incorporated into a multiplication and rooting medium for one or more passages, if they are not phytotoxic to the plant explants. A special problem is presented by cryptic or viable but not cultivable bacteria which can be unable to multiply during many passages, but finally be disclosed in mass. The issue is, therefore, to find and apply tools for detection of different media and/or molecular markers. The above questions are discussed in the present paper based on the literature and results of our own study.
9

Bacteria in plant tissue cultures

61%
Orlikowska T., Zawadzka M.
|
EN
Bacterial contamination is quite frequent in plant tissue cultures although, theoretically, cultures have to be axenic. It is their ubiquity and adaptability to different conditions that enable bacteria to colonize also tissue cultures. Among them there are not only the typical endo- and egzophytic species connected with the plant kingdom, but also those which are common among people. The bacteria species isolated from plant tissue cultures were found to be vitropathogenic (pathogens facultative to in vitro explants), latent (pathogens not virulent in vitro), and cryptic (present in tissues but invisible). Some bacteria produce growth regulators, which can modify the morphogenetic mode of explants. They are all undesirable ones in cultures, but the explants contaminated with pathogenic species should be eliminated obligatorily. Various groups of bacteria, as well as the techniques of detecting, identifying and eliminating them, are briefly described.
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