title: Ð Ð°Ð·ÑÐ°Ð±Ð¾ÑÐºÐ° ÐÐÐ-ÐºÐ¾Ð½ÑÑÑÑÐºÑÐ¸Ð¸ Ð´Ð»Ñ Ñ Ð°ÑÐ°ÐºÑÐµÑÐ¸Ð·Ð°ÑÐ¸Ð¸ Ð¼ÐµÑÐ¾Ð´Ð¾Ð¼ Ð¾Ð¿ÑÐ¸ÑÐµÑÐºÐ¾Ð³Ð¾ Ð·Ð°Ñ Ð²Ð°ÑÐ° Ð¼ÐµÑ Ð°Ð½Ð¸ÑÐµÑÐºÐ¾Ð¹ ÑÑÐ°Ð±Ð¸Ð»ÑÐ½Ð¾ÑÑÐ¸ ÑÐµÐºÐ¾Ð½ÑÑÑÑÐ¸ÑÐ¾Ð²Ð°Ð½Ð½ÑÑ Ð½ÑÐºÐ»ÐµÐ¾ÑÐ¾Ð¼Ð½ÑÑ ÑÐ°ÑÑÐ¸Ñ
title: Ð²ÑÐ¿ÑÑÐºÐ½Ð°Ñ ÐºÐ²Ð°Ð»Ð¸ÑÐ¸ÐºÐ°ÑÐ¸Ð¾Ð½Ð½Ð°Ñ ÑÐ°Ð±Ð¾ÑÐ° Ð±Ð°ÐºÐ°Ð»Ð°Ð²ÑÐ°
titleType: Subtitle
title: Development of a DNA construct for characterization by optical capture of the mechanical stability of reconstituted nucleosome particles
Creators: name: ÐÐ¸Ð½Ð½Ð¸Ðº, ÐÐ¸ÐºÑÐ¾Ñ
Publisher: name: Ð¡Ð°Ð½ÐºÑ-ÐÐµÑÐµÑÐ±ÑÑÐ³ÑÐºÐ¸Ð¹ Ð¿Ð¾Ð»Ð¸ÑÐµÑÐ½Ð¸ÑÐµÑÐºÐ¸Ð¹ ÑÐ½Ð¸Ð²ÐµÑÑÐ¸ÑÐµÑ ÐÐµÑÑÐ° ÐÐµÐ»Ð¸ÐºÐ¾Ð³Ð¾
Year: 2024
Subjects: Ð½ÑÐºÐ»ÐµÐ¾ÑÐ¾Ð¼Ð°
ÑÐµÐ¼Ð¾Ð´ÐµÐ»Ð»Ð¸ÑÐ¾Ð²Ð°Ð½Ð¸Ðµ Ñ ÑÐ¾Ð¼Ð°ÑÐ¸Ð½Ð°
Ð¾Ð´Ð½Ð¾Ð¼Ð¾Ð»ÐµÐºÑÐ»ÑÑÐ½Ð°Ñ Ð±Ð¸Ð¾ÑÐ¸Ð·Ð¸ÐºÐ°
Ð¾Ð¿ÑÐ¸ÑÐµÑÐºÐ¸Ð¹ Ð¿Ð¸Ð½ÑÐµÑ
ÑÐ±Ð¾ÑÐºÐ° ÐÐ¸Ð±ÑÐ¾Ð½Ð°
nucleosome
chromatin remodeling
single-molecule biophysics
optical tweezers
Gibson assembly

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description: In this thesis we describe the DNA preparation protocol for characterization of nucleosome stability at the single-molecule level. The preparation of the DNA construct involved the insertion of the â603â nucleosome positioning sequence into 11.3 kbp plasmid vector using Gibson Assembly cloning, transformation and extraction of the fragment from cell culture after growing sufficient amount of biomass. The final DNA construct was used to reconstitute nucleosomes by gradient salt dialysis with descending salt concentration. The reconstitution of nucleosomes was confirmed by measuring force-extension behavior of individual nucleosomes using dual trap optical tweezers. The obtained results could be further used to study the interaction of nucleosomes with regulatory proteins at single-molecule level.