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Protein-nanodiamond conjugates: the preparation and characterization
Šlegerová, Jitka ; Konvalinka, Jan (advisor) ; Vaněk, Ondřej (referee)
Nanodiamonds represent a member of vast group of carbon nanomaterials. They are characterized by chemical stability, biocompatibility, low toxicity and possibility of surface chemical modification that enables further conjugation of biomolecules, namely proteins. The most important property of nanodiamonds is easily detectable fluorescence resistant towards photobleaching. This luminescence is produced by so called (N-V) centers that can be formed by irradiation of nanodiamonds using high energy beams. Fluorescence of nanodiamonds is useful for construction of fluorescent labels and probes, cellular targeting and internalization, controlled drug delivery as well as enzyme immobilization. Two main connection modes are suitable for biomolecules' attachment: non-covalent and covalent. The first one has been utilized for decoration of nanodiamonds by various proteins, e.g. lysozyme, cytochrome c, neurotoxin or antigen. The second possibility has been demonstrated also on various proteins and, furthermore, on glycoproteins, oligonucleotides, vitamins or growth factor. During the immobilization of biomolecule on the nanoparticle surface it is crucial to keep its function, for example the catalytic activity in enzymes. For such purposes, the oxime ligation (connection of aldehyde presented on biomolecule...
Direct enzymatic synthesis of aldehyde-functionalized DNA and its conjugation with hydrazines and amines
Raindlová, Veronika ; Hocek, Michal
A new simple methodology for DNA conjugation or staining was developed. 2′-Deoxyribonucleoside triphosphates (dNTPs) bearing reactive aldehyde group were prepared by one-step Suzuki cross-coupling reaction of halogenated dNTPs with boronic acid. These modified dNTPs were enzymatically incorporated into DNA by primer extension (PEX) or amplified by polymerase chain reaction (PCR) using different DNA polymerases. The followup reaction between aldehyde-modified PCR products and hydrazine derivatives gave coloured DNA conjugated hydrazones. This methodology was also used for further conjugations of aldehyde-modified 2′-deoxyribonucleoside monophosphates (dNMPs) with amines by reductive amination.

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