Protocol to quantify the rate of RNA polymerase II elongation with MS2/MCP- and PP7/PCP-based live-imaging systems in early Drosophila embryos

STAR Protoc. 2024 Jun 21;5(2):103099. doi: 10.1016/j.xpro.2024.103099. Epub 2024 May 31.

Abstract

The MS2-PP7 two-color live-imaging system provides insights into the spatiotemporal dynamics of nascent transcripts at tagged loci. Here, we present a protocol to quantitatively measure the rate of RNA polymerase II elongation for each actively transcribing nucleus in living Drosophila embryos. The elongation rate is calculated by measuring the effective distance and the time elapsed between MS2 and PP7 trajectories. We describe steps for preparing embryo samples, performing live imaging, and measuring the elongation rate. For complete details on the use and execution of this protocol, please refer to Keller et al.1.

Keywords: Developmental biology; Gene Expression; Model Organisms.

MeSH terms

  • Animals
  • Drosophila / embryology
  • Drosophila / metabolism
  • Drosophila Proteins / genetics
  • Drosophila Proteins / metabolism
  • Drosophila melanogaster / embryology
  • Drosophila melanogaster / genetics
  • Drosophila melanogaster / metabolism
  • Embryo, Nonmammalian* / metabolism
  • RNA Polymerase II* / genetics
  • RNA Polymerase II* / metabolism

Substances

  • RNA Polymerase II
  • Drosophila Proteins