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We introduce a new genosensing approach employing CTAB (cetyltrimethylammonium bromide)-coated positively charged colloidal gold nanoparticles (GNPs) to detect target DNA sequences by using absorption spectroscopy and dynamic light scattering. The approach is compared with a previously reported method employing unmodified CTAB-coated gold nanorods (GNRs). Both approaches are based on the observation that whereas the addition of probe and target ssDNA to CTAB-coated particles results in particle aggregation, no aggregation is observed after addition of probe and nontarget DNA sequences. Our goal was to compare the feasibility and sensitivity of both methods. A 21-mer ssDNA from the human immunodeficiency virus type 1 HIV-1 U5 long terminal repeat (LTR) sequence and a 23-mer ssDNA from the Bacillus anthracis cryptic protein and protective antigen precursor (pagA) genes were used as ssDNA models. In the case of GNRs, unexpectedly, the colorimetric test failed with perfect cigar-like particles but could be performed with dumbbell and dog-bone rods. By contrast, our approach with cationic CTAB-coated GNPs is easy to implement and possesses excellent feasibility with retention of comparable sensitivity--a 0.1 nM concentration of target cDNA can be detected with the naked eye and 10 pM by dynamic light scattering (DLS) measurements. The specificity of our method is illustrated by successful DLS detection of one-three base mismatches in cDNA sequences for both DNA models. These results suggest that the cationic GNPs and DLS can be used for genosensing under optimal DNA hybridization conditions without any chemical modifications of the particle surface with ssDNA molecules and signal amplification. Finally, we discuss a more than two-three-order difference in the reported estimations of the detection sensitivity of colorimetric methods (0.1 to 10-100 pM) to show that the existing aggregation models are inconsistent with the detection limits of about 0.1-1 pM DNA and that other explanations should be developed. [ABSTRACT FROM AUTHOR]

Improving the nutritional value of grain sorghum, a drought- and heat-tolerant grain crop, is an important task in the context of global warming. One of the reasons for the low nutritional value of sorghum grain is the resistance of its storage proteins (kafirins) to proteolytic digestion, which is due, among other things, to the structural organization of protein bodies, in which γ-kafirin, the most resistant to proteases, is located on the periphery, encapsulating more easily digested α-kafirins. The introduction of genetic constructs capable of inducing RNA silencing of the γ-kafirin (gKAF1) gene opens up prospects for solving this problem. Using Agrobacterium-mediated genetic transformation of immature embryos of the grain sorghum cv. Avans we have obtained a mutant with improved digestibility of endosperm proteins (up to 92 %) carrying a genetic construct for RNA silencing of the gKAF1 gene. The goal of this work was to study the stability of inheritance of the introduced genetic construct in T2-T4 generations, to identify the number of its copies, as well as to trace the manifestation of agronomically valuable traits in the offspring of the mutant. The mutant lines were grown in experimental plots in three randomized blocks. The studied lines were characterized by improved digestibility of kafirins, a modified type of endosperm, completely or partially devoid of the vitreous layer, an increased percentage of lysine (by 75 %), reduced plant height, peduncle length, 1000-grains weight, and grain yield from the panicle. In T2, a line with monogenic control of GA resistance was selected. qPCR analysis showed that in different T3 and T4 plants, the genetic construct was present in 2-4 copies. In T3, a line with a high digestibility of endosperm proteins (81 %) and a minimal decrease in agronomically valuable traits (by 5-7 %) was selected., Competing Interests: The authors declare no conflict of interest., (Copyright © AUTHORS.)

Surface-enhanced Raman scattering (SERS) has proven to be a promising technique for different types of imaging including preoperative and intraoperative in vivo tumor visualization. However, the strong scattering of the turbid tissue limits its use in subcutaneous areas. In this article, we used an optical clearing technique to improve the SERS signal from a subcutaneous tumor phantom. The phantom is a 2 mm sphere of calcium alginate with incorporated petal-like gap-enhanced Raman tags. The use of optical clearing increases the SERS signal target-to-background ratio for 5 times and allow to decrease the total imaging time for at least 10 times. In addition, SERS imaging assisted with optical clearing made it possible to more precisely determine the shape and boundaries of the implanted phantom. The combination of optical clearing and SERS is a promising strategy for the clinical imaging of subcutaneous objects that are usually shielded by dermal tissue., (© 2021 Wiley-VCH GmbH.)

Nowadays, tissue engineering is one of the most promising approaches for the regeneration of various tissues and organs, including the cornea. However, the inability of biomaterial scaffolds to successfully integrate into the environment of surrounding tissues is one of the main challenges that sufficiently limits the restoration of damaged corneal tissues. Thus, the modulation of molecular and cellular mechanisms is important and necessary for successful graft integration and long-term survival. The dynamics of molecular interactions affecting the site of injury will determine the corneal transplantation efficacy and the post-surgery clinical outcome. The interactions between biomaterial surfaces, cells and their microenvironment can regulate cell behavior and alter their physiology and signaling pathways. Nanotechnology is an advantageous tool for the current understanding, coordination, and directed regulation of molecular cell-transplant interactions on behalf of the healing of corneal wounds. Therefore, the use of various nanotechnological strategies will provide new solutions to the problem of corneal allograft rejection, by modulating and regulating host-graft interaction dynamics towards proper integration and long-term functionality of the transplant.

The recently developed laser-induced cell transfection mediated by Au nanoparticles is a promising alternative to the well-established lipid-based transfection or to electroporation. Optoporation is based on the laser plasmonic heating of nanoparticles located near the cell membrane. However, the uncontrollable cell damage from intense laser pulses and from random attachment of nanoparticles may be crucial for transfection. We present a novel plasmonic optoporation technique that uses Au nanostar layers immobilized in culture microplate wells. HeLa cells were grown directly on Au nanostar layers, after which they were subjected to continuous-wave 808 nm laser irradiation. An Au monolayer density ~15 μg/cm 2 and an absorbed energy of about 15 to 30 J were found to be optimal for optoporation. Propidium iodide molecules were used as model penetrating agent. The transfection efficiency evaluated using fluorescence microscopy for HeLa cells transfected with pGFP under optimized optoporation conditions (95% ± 5%) was similar to the efficiency of TurboFect. The technique's efficiency (295 ± 10 relative light units, RLU), demonstrated by transfecting HeLa cells with the pCMV-GLuc 2 control plasmid, was greater than that obtained by transfection of HeLa cells with the TurboFect agent (220 ± 10 RLU). The cell viability in plasmonic optoporation (92% ± 7%), too, was greater than that in transfection with TurboFect (75% ± 7%)., (© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Au, Ag, Se, and Si nanoparticles were synthesized from aqueous solutions of HAuCl 4 , AgNO 3 , Na 2 SeO 3 , and Na 2 SiO 3 with extra- and intracellular extracts from the xylotrophic basidiomycetes Pleurotus ostreatus , Lentinus edodes , Ganoderma lucidum , and Grifola frondosa . The shape, size, and aggregation properties of the nanoparticles depended both on the fungal species and on the extract type. The bioreduction of the metal-containing compounds and the formation rate of Au and Ag nanoparticles depended directly on the phenol oxidase activity of the fungal extracts used. The biofabrication of Se and Si nanoparticles did not depend on phenol oxidase activity. When we used mycelial extracts from different fungal morphological structures, we succeeded in obtaining nanoparticles of differing shapes and sizes. The cytotoxicity of the noble metal nanoparticles, which are widely used in biomedicine, was evaluated on the HeLa and Vero cell lines. The cytotoxicity of the Au nanoparticles was negligible in a broad concentration range (1-100 µg/mL), whereas the Ag nanoparticles were nontoxic only when used between 1 and 10 µg/mL., Competing Interests: The authors declare there are no competing interests.

Gold nanoparticles (GNPs) and GNP-based multifunctional nanocomposites are the subject of intensive studies and biomedical applications. This minireview summarizes our recent efforts in analytical and theranostic applications of engineered GNPs and nanocomposites by using plasmonic properties of GNPs and various optical techniques. Specifically, we consider analytical biosensing; visualization and bioimaging of bacterial, mammalian, and plant cells; photodynamic treatment of pathogenic bacteria; and photothermal therapy of xenografted tumors. In addition to recently published reports, we discuss new data on dot immunoassay diagnostics of mycobacteria, multiplexed immunoelectron microscopy analysis of Azospirillum brasilense, materno-embryonic transfer of GNPs in pregnant rats, and combined photodynamic and photothermal treatment of rat xenografted tumors with gold nanorods covered by a mesoporous silica shell doped with hematoporphyrin.