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Chemia bioortogonalna – użyteczne narzędzie badania procesów wewnątrzkomórkowych

Latos Krystian

Wiadomości Chemiczne

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2022

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[Z] 76, 1-2

79--95

EN

Bioorthogonal chemistry is a rapidly developing field of science operating on the border of chemistry and biology. Its initial goal was to study metabolism and imaging using fluorescently labelled compounds. Due to recent advances, bioorthogonal chemistry can also be used to engineer therapeutic bioconjugates. By using a combination of bioconjugation and advanced omics techniques, it is possible to study and modify complex interactions inside living cells. In the relatively short time since its introduction, bioorthogonal chemistry has found many applications. In nucleic acid research, it is used for labelling, e.g. with biotin, to facilitate detection, immobilization, and purification. Additionally, thanks to the use of fluorescent nucleoside analogues, it can be used to study the interaction and dynamics of nucleic acids. For the study of proteins, bioorthogonal chemistry is an invaluable tool for studying conformation, as well as intramolecular and intermolecular interactions. Using techniques such as PET and FRET it is possible to take a closer look at the structure of proteins, which has a significant impact on their functionality. By using biarsenical dyes, interactions between proteins are tracked. This is used in the study of protein aggregation in diseases such as Alzheimer's, Huntington's, and prion diseases. Thanks to this, it becomes possible to understand the mechanism and pathology of these diseases. In biosensing, the elements of bioorthogonal chemistry have been used in a variety of tests and imaging methods. In the end, methods for testing glycan are presented. The advantage of bioorthogonal methods is that they allow labelling on the whole cell or lysate. This application in glycoproteomics is extremely important due to the fact that changes in glycosylation occur during disease states.

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Przegląd aktualnie dostępnych technologii sekwencjonowania kwasów nukleinowych

Boczkowska Maja

Laboratorium - Przegląd Ogólnopolski

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2022

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nr specjalny

42--48

PL

Sekwencjonowanie DNA-Seq wykorzystuje się do sekwencjonawania genomów, eksomów, epigenomu, targetowego resekwencjonowania fragmentów genomów, genotypowania. RNA-Seq można podzielić na: sekwencjonowanie całego transkryptomu, mRNA (mRNA-Seq) i małych RNA (smRNA-Seq). Jest wykorzystywane w badaniach z zakresu genomiki funkcjonalnej, różnicowej analiz ekspresji genów, alternatywnego splicingu i analizy wariantów.

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Przegląd aktualnie dostępnych technologii sekwencjonowania kwasów nukleinowych

Boczkowska Maja

Laboratorium - Przegląd Ogólnopolski

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2021

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nr 3

14--21

PL

Sekwencjonowanie DNA-Seq wykorzystuje się do sekwencjonowania genomów, eksomów, epigenomu, targetowego resekwencjonowania fragmentów genomów, genotypowania. RNA-Seq można podzielić na: sekwencjonowanie całego transkryptomu, mRNA (mRNA-Seq) i małych RNA (smRNA-Seq). Jest wykorzystywane w badaniach z zakresu genomiki funkcjonalnej, różnicowej analizy ekspresji genów, alternatywnego splicingu i analizy wariantów.

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Analiza oligonukleotydów antysensownych pierwszej i drugiej generacji za pomocą chromatografii par jonowych

Kaczmarkiewicz Anna, Nuckowski Łukasz, Studzińska Sylwia, Buszewski Bogusław

Analityka : nauka i praktyka

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2019

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nr 2

4--11

PL

Rosnące zainteresowanie badaczy potencjalnymi terapeutykami, jakimi są oligonukleotydy antysensowne, sprawia, że wymagana jest ich czuła analiza na różnych etapach badań klinicznych.

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Oligonukleotydy DNA jako warstwy receptorowe sensorów elektrochemicznych

Górski Ł., Ziółkowski R., Bala A., Jarczewska M., Malinowska E.

Wiadomości Chemiczne

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2015

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[Z] 69, 5-6

325--336

EN

The need for elaboration of analytical devices of small dimensions and the accessibility of novel nanomaterials caused the increase in the number of publications referring to the development of biosensors. DNA-based biosensors are of special interest and they were primarily used for the determination of a specific sequence which is crucial in the detection of cancer, genetic mutations, pathogens, as well as analysis of modified food. Interestingly, they could be also applied for the detection of other analytes including heavy metal ions, especially in connection with electrochemical techniques. It should be noted that the design of DNA biosensor concerns not only the development of transducer, but also careful preparation of sensing layer and the choice of the method of analytical signal generation. Selectivity is one of the essential parameter of the biosensor that determines its utility, particularly in real samples of complex matrices. In case of DNA sensors dedicated for the detection of complementary sequence, high selectivity is provided by the hybridization process. A pronounced specificity of sensing layer-analyte interaction can be also achieved with the use of functional nucleic acids - aptamers, which change their conformation upon binding an analyte. Herein, DNA-modified electrodes were firstly used for the detection of uranyl ions, as they exhibit high affinity towards phosphate moieties of nuclec acids. It was shown that UO2 2+ interacts with sensing layer independently from the chosen oligonucleotide sequence. Moreover, the influence of Pb2+ was reduced by elimination of adenine, which strongly interacts with lead ions. Another oligonucleotide-based sensor was developed for detection of mercury ions. The results indicate that Hg2+ concentration can be determined only with the use of sequence containing 100% thymine residues. Oligonucleotide-based sensor with receptor layer containing aptamers was elaborated for the detection of Pb2+ ions. In the presence of lead cations, an aptamer probe forms a G-quadruplex structure, a proposed biosensor could be characterized with selectivity towards Pb2+ The performance of DNA-based sensors for UO2 2+, Hg2+ and Pb2+ ions was optimized and addressed the choice of the manner of analytical signal generation, the influence of electrode modification with blocking agent, sensitivity dependence on the oligonucleotide sequence and the possibility of regeneration of sensing layer. Finally, the utility of proposed DNA sensors was tested by analysis in real samples.

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Amplifikacja kwasów nukleinowych w warunkach izotermicznych - cz. I

Hołysz M., Hołysz H., Burzyński A.

Laboratorium - Przegląd Ogólnopolski

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2015

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nr 9-10

59--64

PL

Amplifikacja kwasów nukleinowych w warunkach izotermicznych stanowi ciekawą alternatywę wobec technik tradycyjnie wykorzystywanych w badaniach naukowych i w diagnostyce. W ciągu ostatnich kilkunastu lat opracowano wiele różnych metod powielania DNA i RNA w stałej temperaturze. Techniki te nie ustępują pod względem wielu parametrów klasycznej reakcji PCR, nawet technice real-time PCR, a niekiedy nawet je przewyższają. Cele niniejszego artykułu to: prezentacja technik izotermicznej amplifikacji kwasów nukleinowych, wyjaśnienie zasady ich działania oraz prezentacja produktów diagnostycznych działających w oparciu o te techniki.

EN

Amplification of nucleic acids in isothermal conditions is an interesting alternative for techniques traditionally used in research and diagnostics. For the last several years many different methods of DNA and RNA replication in constant temperature have been developed. These techniques does not leg behind, in terms of many parameters, the classical PCR, and even real-time PCR technique, and sometimes they are even better. The aim of the article is to present the techniques of isothermal amplification of nucleic acids, explain their basic principles and present diagnostic products operating based on these techniques.

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Dializa równowagowa. Metoda badania selektywności oddziaływań ligand.DNA

Czerwińska I., Głuszyńska A., Juskowiak B.

Wiadomości Chemiczne

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2010

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[Z] 64, 1-2

105-122

EN

Equilibrium or competition dialysis is a powerful tool for binding study of ligands that are expected to bind to nucleic acids with selectivity related to their structure or sequence. In the equilibrium dialysis experiment, a set of nucleic acid samples that differ in structure and sequence is dialyzed against a test ligand solution. After equilibration, the concentration of ligand bound to each structure or sequence is determined by UV-Vis absorption or fluorescence spectroscopy in each dialysis unit. Since all nucleic acid samples are in equilibrium with the same free ligand concentration, the amount of bound ligand is directly related to the ligand binding affinity. Thus, equilibrium provides a direct measure of selectivity and identifies the nucleic acid sample, which is preferred by a particular ligand. We describe here the principles and practice of the method. Examples of an application of the method are limited to the discovery of small molecules that selectively recognize the unique structural features of G-quadruplexes. There are proofs for important functional roles of G-quadruplex structures in biology (maintenance of telomeres, transcriptional regulation, and modulation of mRNA translation). G-quadruplex DNA can exist in a variety of structural forms that may possess numerous potential binding sites for small molecules. Therefore equilibrium dialysis provides a useful tool for discovery of new mall-molecule therapeutic agents targeting G-quadruplexes.

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Krótkie cząsteczki kwasów nukleinowych jako terapeutyki

Sipa K.

Laboratorium - Przegląd Ogólnopolski

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2004

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nr 4

38--42

PL

Pół wieku po opisaniu przez Watsona i Cricka struktury DNA nasze rozumienie procesów leżących u podstaw życia jest coraz pełniejsze, ale wciąż dalekie od kompletnego. Uniwersalne zasady przepływu informacji genetycznej w komórce są uzupełniane przez odkrycia rzucające światło na procesy regulacji ekspresji genów, w których doniosłą rolę pełnią cząsteczki RNA – drugiego, starszego ewolucyjnie kwasu nukleinowego.

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Biowskaźniki DNA : wykorzystanie kwasów nukleinowych w analityce

Filipiak M.

Analityka : nauka i praktyka

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2004

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nr 3

10-13