| Title: |
Rapid profiling of DNA replication dynamics using mass spectrometry–based analysis of nascent DNA |
| Authors: |
Ashour, Mohamed E.; Byrum, Andrea K.; Meroni, Alice; Xia, Jun; Singh, Saurabh; Galletto, Roberto; Rosenberg, Susan M.; Vindigni, Alessandro; Mosammaparast, Nima |
| Contributors: |
American Italian Cancer Foundation; Alvin J. Siteman Cancer Center; Foundation for Barnes-Jewish Hospital; Cancer Frontier Fund; U.S. Department of Defense; State of Nebraska; National Institutes of Health; American Cancer Society; Centene Corporation; Barnard Foundation |
| Source: |
Journal of Cell Biology ; volume 222, issue 4 ; ISSN 0021-9525 1540-8140 |
| Publisher Information: |
Rockefeller University Press |
| Publication Year: |
2023 |
| Description: |
The primary method for probing DNA replication dynamics is DNA fiber analysis, which utilizes thymidine analog incorporation into nascent DNA, followed by immunofluorescent microscopy of DNA fibers. Besides being time-consuming and prone to experimenter bias, it is not suitable for studying DNA replication dynamics in mitochondria or bacteria, nor is it adaptable for higher-throughput analysis. Here, we present mass spectrometry–based analysis of nascent DNA (MS-BAND) as a rapid, unbiased, quantitative alternative to DNA fiber analysis. In this method, incorporation of thymidine analogs is quantified from DNA using triple quadrupole tandem mass spectrometry. MS-BAND accurately detects DNA replication alterations in both the nucleus and mitochondria of human cells, as well as bacteria. The high-throughput capability of MS-BAND captured replication alterations in an E. coli DNA damage-inducing gene library. Therefore, MS-BAND may serve as an alternative to the DNA fiber technique, with potential for high-throughput analysis of replication dynamics in diverse model systems. |
| Document Type: |
article in journal/newspaper |
| Language: |
English |
| DOI: |
10.1083/jcb.202207121 |
| DOI: |
10.1083/jcb.202207121/1447856/jcb_202207121.pdf |
| Availability: |
https://doi.org/10.1083/jcb.202207121; https://rupress.org/jcb/article-pdf/doi/10.1083/jcb.202207121/1447856/jcb_202207121.pdf |
| Rights: |
http://www.rupress.org/terms/ ; https://creativecommons.org/licenses/by-nc-sa/4.0/ |
| Accession Number: |
edsbas.1D33663D |
| Database: |
BASE |