2022
DOI: 10.1101/2022.06.22.497140
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Extracting and characterizing protein-free megabasepair DNA for in vitro experiments

Abstract: Chromosome structure and function is studied in cells using imaging and chromosome-conformation-based methods as well as in vitro with a range of single-molecule techniques. Here we present a method to obtain genome-size (megabasepair length) deproteinated DNA for in vitro studies, which provides DNA substrates that are two orders of magnitude longer than typically studied in single-molecule experiments. We isolated chromosomes from bacterial cells and enzymatically digested the native proteins. Mass spectrome… Show more

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“…The chromosome isolation protocol was adapted from different works. , E. coli eGFP16 was grown in 25 mL of liquid LB medium until an OD600 of 1.2–1.5, and cells were pelleted.…”
Section: Methodsmentioning
confidence: 99%
See 1 more Smart Citation
“…The chromosome isolation protocol was adapted from different works. , E. coli eGFP16 was grown in 25 mL of liquid LB medium until an OD600 of 1.2–1.5, and cells were pelleted.…”
Section: Methodsmentioning
confidence: 99%
“…The chromosome isolation protocol was adapted from different works. 25 , 47 51 E. coli eGFP16 was grown in 25 mL of liquid LB medium until an OD600 of 1.2–1.5, and cells were pelleted. Pellets were recovered with 200 μL of TAE buffer (40 mM Tris-Acetate, 1 mM EDTA, pH 7) and stored in microcentrifuge tubes at −80 °C for further use.…”
Section: Methodsmentioning
confidence: 99%