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Atomic Force Microscope for Topographic Studies
Published in Yuri L. Lyubchenko, An Introduction to Single Molecule Biophysics, 2017
The AFM time-lapse experiments directly illustrate a highly dynamic feature of mononucleosomes, suggesting that nucleosomes are rather unstable. This finding is in line with direct measurements of the NCP stability in diluted solutions in experiments in which the ratio of intact nucleosomes as a function of time was measured by AFM imaging (Menshikova et al. 2011). At the same time, we need to take into account the effect of the surface used for the AFM experiment. Although APS-mica has a low charge density and has little effect on the ion distribution near the surface (Lyubchenko and Shlyakhtenko 1997), the interaction of DNA with the surface favors the unwrapping process. It was hypothesized by Miyagi et al. (2011) that electrostatics can play a role in chromatin unfolding by remodeling factors. Specifically it was posited that the surface of chromatin remodeling factors accommodating the nucleosome would be positively charged to facilitate nucleosome unfolding, and a study from the Kornberg lab (Lorch et al. 2010) supports this hypothesis. The effect of local surface properties on nucleosome dynamics can be generalized to the chromatin level. The interaction of chromatin with nuclear membranes and other components of the nuclear matrix are important factors that modulate chromatin structure and function. Designing the surfaces with various characteristics would mimic the membrane structure, and the AFM as a topographic technique is an ideal method to characterize these interactions.
Development of an improved lentiviral based vector system for the stable expression of monoclonal antibody in CHO cells
Published in Preparative Biochemistry and Biotechnology, 2019
Omid Mohammadian, Masoumeh Rajabibazl, Es’hagh Pourmaleki, Hadi Bayat, Roshanak Ahani, Azam Rahimpour
Utilization of the cis-acting elements including SAR/MARs in the expression construct has proven to reduce the chromosomal position effects. SARs are able to attach the DNA to the nuclear matrix thereby generating an active chromatin domain. It has also been shown that SARs can recruit transcription enhancing trans-factors.[34] Also SAR characteristic motifs play an important role in overcoming transgene silencing by affecting histone demethylation and acetylation.[35] Various types of SAR elements have been identified from different sources including the human ß interferon SAR, human ß globin SAR, and chicken lysozyme 5′ SAR.[36] Human beta interferon SAR (IFN-SAR) previously shown to increase the stability and long-term transgene expression from plasmid-based vectors in CHO cells.[21] In addition, the positive effects of this element on transgene expression from retoviral and lentiviral vectors in primary human CD34+ cord blood cells and hematopoietic stem cells (HSCs) have been shown.[28,37]
Nonviral gene delivery using PAMAM dendrimer conjugated with the nuclear localization signal peptide derived from human papillomavirus type 11 E2 protein
Published in Journal of Biomaterials Science, Polymer Edition, 2021
Jeil Lee, Yong-Eun Kwon, Jaegi Kim, Dong Woon Kim, Hwanuk Guim, Jehyeong Yeon, Jin-Cheol Kim, Joon Sig Choi
We hypothesized that PAMAM derivatives conjugated with NLS peptides would induce perinuclear accumulation of polyplexes. Although the gene delivery mechanism of HPV type 11 has not been established, the central hinge domain of the viral E2 protein is known to deliver viral DNA stably by combining with the nuclear matrix and matrix-associated proteins. The RKRAR sequence containing the central hinge domain is a key factor in the nuclear localization of viral DNA [16].